-
This invention was made with Government Support under Grant No.
DK52798, awarded by the National Institutes of Health. The Government has certain
rights to this invention.
INTRODUCTION
Technical Field
-
This invention relates to thyroid hormone agonists and antagonists, methods of
using such compounds, and pharmaceutical compositions containing them. The
invention also relates to methods of preparing such compounds.
Background
-
Nuclear receptors represent a superfamily of proteins that specifically bind a
physiologically relevant small molecule, such as hormone or vitamin. As a result of a
molecule binding to a nuclear receptor, the nuclear receptor changes the ability of a
cell to transcribe DNA, i.e. nuclear receptors modulate the transcription of DNA,
although they may have transcription independent actions. Unlike integral membrane
receptors and membrane associated receptors, the nuclear receptors reside in either the
cytoplasm or nucleus of eukaryotic cells. Thus, nuclear receptors comprise a class of
intracellular, soluble ligand-regulated transcription factors.
-
Nuclear receptors include receptors for thyroid hormones. Thyroid hormones
promote normal growth and development and control an extraordinary number of
regulatory functions in mammals. They regulate fetal development, cholesterol
metabolism, the level of obesity, free radical formation, intestinal and cardiovascular
functions, and bone and calcium metabolism. In current medical practice, thyroid
hormones are used mostly for replacement therapy for humans with hypothyroidism,
and to suppress the pituitary gland stimulation of the thyroid gland in patients with
thyroid nodules or cancer. However, these hormones cannot be administered in high
doses because of significant side effects, mainly on the heart.
-
There are two major subtypes of the thyroid hormone receptor ("TR"), TR( and
TR(, and they are expressed from two different genes. Preliminary experiments
indicate that the ( and ( subtypes are differentially expressed in various tissues.
-
It is desirable to produce thyroid hormone agonists and antagonists that are
selective for TR( and TR(. Surprisingly, a small class of halogen-free thyroid
hormone agonists has been discovered, which are highly selective for the TR( subtype
with high binding affinity, and are described in U.S. Patent No. 5,883,294, the
complete disclosure of which is hereby incorporated by reference. Another previous
disclosure of interest is U.S. Patent Application 08/764,870, filed December 13, 1995,
the complete disclosure of which is hereby incorporated by reference.
-
Although antagonist ligands have been developed for a number of nuclear
receptors, there is currently no reported high-affinity antagonist for the TR.
Examination of known nuclear receptor antagonist ligands reveals that these
compounds structurally resemble their agonist counterparts but contain a large (> 8
carbon atom) extension group attached to the middle of the molecule, Ribeiro, et al.,
Recent Prog. Horm. Res. 53:351-394 (1998).
-
A high affinity TR(-selective agonist ligand, designated GC-1, was recently
prepared and characterized, Chiellini, et al., Chemistry & Biology 5:299-306 (1998).
GC-1 contains several structural differences from 3,5,3'-triodo-L-thyronine ("T3"), the
major active form of thyroid hormone. In particular, the methylene unit bridging the
two phenyl rings introduces a new derivatizable position in the middle of the molecule
which is unavailable in the natural ligand where an ether oxygen joins the rings.
-
It would be highly desirable to design a route for the efficient derivatization of
the bridging carbon present on the GC-1 family of compounds with a panel of
nucleophiles. This has been successful in the preparation of semi-synthetic rapamycin
derivatives by using an SN1 reaction, Luengo, et al., J. Org. Chem., 59:6512 (1994)
and Luengo, et al., Chem. Biol., 2:471 (1995). The resulting GC-1 derivatives can
then be converted to thyromimetics using the same chemistry that was established for
the synthesis of GC-1. It is expected that these thyromimetics, which contain an
extension group attached to the middle of a TR agonist, would be high-affinity
antagonists of TR.
SUMMARY OF THE INVENTION
-
-
The present invention relates to compounds of Formula I:
wherein:
- n is 1, 2 or 3;
- R1 is C112alkyl, C3-12alkanol, C2-6alkenyl, C5-12alkenol, heterocyclo, aryl
substituted with at least one electron-donating group, -OR2 or SR2, where R2 is
C1-12alkyl or aryl, or AC(O)NR12R13, where A is C215alkyl or C415alkenyl and R12 and
R13 are C16alkyl;
- R3 and R5 are methyl;
- R4 is hydrogen, C16alkyl or cycloalkyl;
- R6 and R9 are hydrogen or C16alkyl;
- R7 and R8 are independently hydrogen, halogen, C16alkyl, optionally
substituted phenyl, optionally substituted benzyl, or heteroaryl; with the proviso that
R7 and R8 cannot both be hydrogen;
- R10 is hydrogen, C16alkyl, cycloalkyl, or acyl; and
- R11 is hydrogen, C16alkyl, or cycloalkyl;
and the pharmaceutically acceptable salts thereof. -
-
In a second aspect, the invention relates to a method of treatment of mammals
having a disease state that is treatable by thyroid hormones, comprising administering
a therapeutically effective dose of a compound of Formula I.
-
In a third aspect, the invention relates to a pharmaceutical composition
containing a therapeutically effective amount of a compound of Formula I admixed
with at least one pharmaceutically acceptable excipient.
-
In a fourth aspect, the invention relates to processes for preparing the
compounds of Formula I.
-
The invention also relates to compounds of Formula II:
where Y is -OT or -O(CH2)
nCO
2C
1-6alkyl; X and T are protecting groups, and n, R
1
and R
3-R
9 are as defined above.
-
In another aspect, the invention relates to the use of a compound of Formula II
as an intermediate in the process for preparing the compounds of Formula I.
-
The invention also relates to compounds of Formula III:
where Y' is -OT' or -O(CH
2)
nCO
2C
1-6alkyl; X' and T' are protecting groups, and at least
one of said protecting groups is a silyl containing protecting group; Z' is a leaving
group and n and R
3-R
9 are as defined above.
-
In another aspect, the invention relates to the use of a compound of Formula III
as an intermediate in the process for preparing the compounds of Formula I.
-
In yet another aspect, the invention pertains to the processes for preparing
compounds of Formulas II and III.
DETAILED DESCRIPTION OF THE INVENTION
-
The invention relates to a family of compounds useful for treating a disease
state that is treatable by thyroid hormones. These compounds have the Formulas I, II
and III as set forth below.
-
The present invention relates to compounds of Formula I:
wherein:
- n is 1, 2 or 3;
- R1 is C1/2alkyl, C3-12alkanol, C2-6alkenyl, C5-12alkenol, heterocyclo, aryl
substituted with at least one electron-donating group, -OR2 or SR2, where R2 is
C1-12alkyl or aryl, or AC(O)NR12R13, where A is C15alkyl or C415alkenyl and R12 and
R13 are C16alkyl;
- R3 and R5 are methyl;
- R4 is hydrogen, C16alkyl or cycloalkyl;
- R6 and R9 are hydrogen or C16alkyl;
- R7 and R8 are independently hydrogen, halogen, C16alkyl, optionally
substituted phenyl, optionally substituted benzyl, or heteroaryl; with the proviso that
R7 and R8 cannot both be hydrogen;
- R10 is hydrogen, C16alkyl, cycloalkyl, or acyl; and
- R11 is hydrogen, C16alkyl, or cycloalkyl;
and the pharmaceutically acceptable salts thereof.-
-
The invention also relates to compounds of Formula II, which find utility as
intermediates in the process for preparing the compounds of Formula I:
where Y is -OT or -O(CH
2)
nCO
2C
1-6alkyl; X and T are protecting groups, and n, R
1
and R
3-R
9 are as defined above.
-
The invention also relates to compounds of Formula III, which find utility as
intermediates in the process for preparing the compounds of Formula I:
where Y' is -OT' or -O(CH
2)
nCO
2C
1-6alkyl; X' and T' are protecting groups, and at least
one of said protecting groups is a silyl containing protecting group; Z' is a leaving
group and n and R
3-R
9 are as defined above.
Definitions
-
- "Alkyl" means a branched or unbranched saturated monovalent hydrocarbon
radical containing 1 to 20 carbon atoms (C1-20alkyl), more typically C1-12alkyl, such as
methyl, ethyl, propyl, tertbutyl, nhexyl, noctyl and the like. "Lower alkyl" means an
alkyl group containing 1 to 6 carbon atoms, such as methyl, ethyl, propyl, isopropyl,
tertbutyl, butyl, nhexyl and the like, unless otherwise indicated.
- "Cycloalkyl" as used herein means a saturated monovalent monocyclic
hydrocarbon radical containing 312 carbon atoms, such as cyclopropyl, cyclobutyl,
cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl. The radical may be optionally
mono-, di-, or tri-substituted, independently, with alkyl, lower alkyl, cycloalkyl,
hydroxy-lower alkyl, amino-lower alkyl, hydroxyl, thiol, amino, halo, nitro, lower
alkylthio, lower alkoxy, monolower alkylamino, di-lower alkylamino,
hydroxycarbonyl, lower alkoxycarbonyl, hydroxysulfonyl, lower alkoxysulfonyl,
lower alkylsulfonyl, lower alkylsulfinyl, trifluoromethyl, cyano, tetrazoyl, carbamoyl,
lower alkylcarbamoyl, and dilower alkylcarbamoyl.
- "Lower alkoxy" or "C1-6alkoxy" means the group O-(lower alkyl), wherein
lower alkyl is as herein defined.
- "Alkenyl" means an unsaturated branched or straight chain or alkene radical
containing 2 to 12 carbon atoms and containing a double bond. "Lower alkenyl" or
"C2-6alkenyl" refers to an alkenyl radical of 26 carbon atoms and containing a double
bond. The term is further exemplified by such radicals as ethylene and propylene.
- "Alkanol" and "alkenol" are terms used to mean an alkyl or alkenyl group that
is substituted with a hydroxyl group. Accordingly, "C3-12alkanol" is an alkyl group
having 3-12 carbons and an -OH group, while "C5-12alkenol" is an alkenyl group
having 5-12 carbons and an -OH group.
- "Halo" or "halogen" means fluoro, chloro, bromo, or iodo.
-
-
The term "aryl" refers to a monovalent unsaturated aromatic carbocyclic
radical having a single ring (e.g., phenyl) or two rings (e.g., naphthyl or biphenyl),
which can optionally be mono, di or trisubstituted, independently, with OH, COOH,
lower alkyl, lower alkoxy, nitro, amino, alkylamino, dialkylamino, trifluoromethyl
and/orcyano.
-
The term "acyl" refers to the group -C(O)R, where R is lower alkyl or
cycloalkyl, for example acetyl, propionyl, cyclopropionyl, butanoyl, and the like.
-
The term "heteroatom" refers to oxygen, sulfur and nitrogen, unless otherwise
specified.
-
The term "heterocycloalkyl" refers to a cycloalkyl radical, as defined above,
having 1-3 heteroatoms within the ring (e.g., piperidinyl, piperazinyl, pyrrolidinyl,
pyrrolodinonyl, tetrahydrofuranyl, morpholinyl, tetrahydrothiophenyl, and the like).
The term "heteroaryl" refers to an aryl radical, as defined above, having 1-3
heteroatoms within a single ring (e.g., pyridyl, imidazolyl, thiazolyl, pyrimidine,
oxazolyl, and the like). The term "heterocyclo" is used to collectively refer to
heterocycloalkyl and heteroaryl radicals. The heterocyclo radical can optionally be
mono, di or trisubstituted, independently, with OH, COOH, lower alkyl, lower alkoxy,
nitro, amino, alkylamino, dialkylamino, trifluoromethyl and/or cyano. The term
"heterocyclo" also includes instances where an atom of the heterocyclo has been
oxidized, e.g., Noxides, sulfoxides and sulfones.
-
The term "electron-donating group" refers to a substituent which, when bound
to a molecule, is capable of polarizing the molecule such that the electron-donating
group becomes electron poor and positively charged relative to another portion of the
molecule, i.e., it has reduced electron density. Such groups include, by way of
illustration and not limitation, alkoxys such as methoxy, hydroxy, amines, ethers,
thioethers, phosphines, oxyanions, mercaptans, and their anions, sulfides, etc.
Similarly, the term "aryl substituted with at least one electron-donating group" refers
to an aryl group, preferably phenyl, substituted with at least one, and preferably two,
groups that are electron-donating groups.
-
The term "protecting group as used herein means a radical group that is
covalently bonded to a potentially reactive functionality, masking its reactive nature
and thereby preventing undesired side reactions during the course of chemical
synthesis. For example, a trialkyl silyl protecting group can serve to protect a
hydroxyl functionality, etc. A protecting group preferably is easily attached to the
molecule and also has the property that it may be removed selectively at a desired
point in the chemical synthesis, under conditions that do not harm other functional
groups in the molecule, to yield the unmasked chemical functionality. Suitable
protecting groups include lower alkyls such as methyl, and silyl containing protecting
groups such as triisopropylsilyl ("TIPS") and tert-butylmethoxyphenylsilyloxy
("TBMPS").
-
The term "leaving group as used herein means a group of charged or
uncharged atoms that departs during a substitution or displacement reaction. Suitable
leaving groups include hydroxy and lower alkoxys.
-
"Optional" or "optionally" means that the subsequently described event or
circumstance may or may not occur, and that the description includes instances where
said event or circumstance occurs and instances in which it does not. For example,
optionally substituted phenyl indicates either unsubstituted phenyl, or phenyl mono, di
or trisubstituted, independently, with OH, COOH, lower alkyl, lower alkoxy, halo,
nitro, amino, alkylamino, dialkylamino, trifluoromethyl and/or cyano.
-
As used herein, the terms "inert organic solvent" or "inert solvent" mean a
solvent inert under the conditions of the reaction being described in conjunction
therewith. Such solvents include, by way of example and not limitation, benzene,
toluene, acetonitrile, tetrahydrofuran ("THF"), N,N-dimethylformamide ("DMF"),
chloroform ("CHCl3"), methylene chloride (or dichloromethane or "CH2Cl2"), diethyl
ether, ethyl acetate, acetone, methylethyl ketone, methanol, ethanol, propanol,
isopropanol, tert-butanol, dioxane, pyridine, and the like. Unless specified to the
contrary, the solvents used in the reactions of the present invention are inert solvents.
-
"Pharmaceutically acceptable salt" means those salts which retain the
biological effectiveness and properties of the compounds of Formula I, and which are
not biologically or otherwise undesirable. Such salts may be prepared from inorganic
and organic bases. Salts derived from inorganic bases include, but are not limited to,
the sodium, potassium, lithium, ammonium, calcium, and magnesium salts. Salts
derived from organic bases include, but are not limited to, salts of primary, secondary
and tertiary amines, substituted amines including naturally-occurring substituted
amines, and cyclic amines, including isopropylamine, trimethylamine, diethylamine,
triethylamine, tripropylamine, ethanolamine, 2dimethylaminoethanol, tromethamine,
lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine,
ethylenediamine, glucosamine, Nalkylglucamines, theobromine, purines, piperazine,
piperidine, and Nethylpiperidine. It should also be understood that other carboxylic
acid derivatives would be useful in the practice of this invention, for example
carboxylic acid amides, including carboxamides, lower alkyl carboxamides, di(lower
alkyl) carboxamides, and the like.
-
The term "q.s" is used herein to mean adding a quantity sufficient to achieve a
stated function., for example to bring a solution to a desired volume (q.s. to 100 ml) or
to a desired pH (q.s. to pH 4).
-
It should be understood that Formula I as drawn is intended to represent the
racemic form of compounds of Formula I as well as the individual enantiomers and
nonracemic mixtures thereof, although for the sake of clarity only one enantiomer is
shown. The scope of the invention as described and claimed encompasses the racemic
forms of the compounds of Formula I as well as the individual enantiomers and
nonracemic mixtures thereof.
-
The term "treatment" as used herein covers any treatment of a disease in a
mammal, particularly a human, and includes: (i) preventing the disease from occurring
in a subject which may be predisposed to the disease but has not yet been diagnosed as
having it; (ii) inhibiting the disease, i.e. arresting its development; or (iii) relieving the
disease, i.e. causing regression of the disease.
-
The term "disease state which is alleviated by treatment with a thyroid
hormone antagonist" as used herein is intended to cover all disease states which are
generally acknowledged in the art to be usefully treated with thyroid hormone
antagonists in general, and those disease states which have been found to be usefully
treated by the thyroid hormone antagonists of our invention, the compounds of
Formula I. Such disease states include, but are not limited to, hyperthyroidism,
tachycardia, cardiac arrhythmia, Graves disease, and so forth.
-
The term "therapeutically effective amount" refers to that amount which is
sufficient to effect treatment, as defined above, when administered to a mammal in
need of such treatment. The therapeutically effective amount will vary depending on
the subject and disease state being treated, the severity of the affliction and the manner
of administration, and may be determined routinely by one of ordinary skill in the art.
METHODS OF TREATMENT
-
The compounds of Formula I can be useful in medical treatments and their
biological activity can be measured in the following tests:
- (i) the induction of mitochondrial (-glycerophosphate dehydrogenase
(GPDH:EC 1.1.99.5). This assay is particularly useful since in certain species e.g. rats
it is induced specifically by thyroid hormones and thyromimetics in a close-related
manner in responsive tissues e.g. liver, kidney and the heart (Westerfield, W.W.,
Richert, D.A. and Ruegamer, W.R., Endocrinology, 1965, 77, 802). The assay allows
direct measurement in rates of a thyroid hormone-like effect of compounds and in
particular allows measurement of the direct thyroid hormone-like effect on the heart;
- (ii) the elevation of basal metabolic rate as measured by the increase in
whole body oxygen consumption;
- (iii) the stimulation of the rate of beating of atria isolated from animals
previously dosed with thyromimetics;
- (iv) the change in total plasma cholesterol levels as determined using a
cholesterol oxidase kit (for example, the Merck CHOD iodine colourimetric kit);
- (v) the measurement of LDL (low density lipoprotein) and HDL (high
density lipoprotein) cholesterol in lipoprotein fractions separated by
ultracentrifugation; and p (vi) the change in total plasma triglyceride levels as
determined using enzymatic color tests, for example the Merck System GPO-PAP
method.
-
-
The compounds of Formula I can be found to exhibit anti-thyromimetic
activity in these tests, by: (a) binding to thyroid hormone receptors ((,() by standard in
vitro binding assays, such as are well known in the art; (b) influencing the expression
of genes regulated by the thyroid receptor, measure by in vivo or in vitro experiments,
such as are well known in the art.
-
The compounds of Formula I may therefore be used in therapy, in the
treatment of conditions which can be alleviated by compounds which antagonize the
effects of thyroid hormones in certain tissues. For example, compounds of Formula I
which block the effects of the thyroid hormone are indicated in the treatment of
hypothyroidism. Such compounds are also indicated for use as anti-arrhythmic
agents.
-
In therapeutic use the compounds of the present invention are usually
administered in a standard pharmaceutical composition.
-
The present invention therefore provides in a further aspect pharmaceutical
compositions comprising a compound of Formula I or a pharmaceutically acceptable
salt thereof and a pharmaceutically acceptable carrier. Such compositions include
those suitable for oral, parenteral or rectal administration.
PHARMACEUTICAL COMPOSITIONS
-
Compounds of Formula I and their pharmaceutically acceptable salts which are
active when given orally can be formulated as liquids for example syrups, suspensions
or emulsions, tablets, capsules and lozenges.
-
A liquid composition will generally consist of a suspension or solution of the
compound or pharmaceutically acceptable salt in a suitable liquid carrier(s), for
example ethanol, glycerine, sorbitol, non-aqueous solvent such as polyethylene glycol,
oils or water, with a suspending agent, preservative, surfactant, wetting agent,
flavoring or coloring agent. Alternatively, a liquid formulation can be prepared from
a reconstitutable powder.
-
For example a powder containing active compound, suspending agent, sucrose
and a sweetener can be reconstituted with water to form a suspension; and a syrup can
be prepared from a powder containing active ingredient, sucrose and a sweetener.
-
A composition in the form of a tablet can be prepared using any suitable
pharmaceutical carrier(s) routinely used for preparing solid compositions. Examples
of such carriers include magnesium stearate, starch, lactose, sucrose, microcrystalline
cellulose and binders, for example polyvinylpyrrolidone. The tablet can also be
provided with a color film coating, or color included as part of the carrier(s). In
addition, active compound can be formulated in a controlled release dosage form as a
tablet comprising a hydrophilic or hydrophobic matrix.
-
A composition in the form of a capsule can be prepared using routine
encapsulation procedures, for example by incorporation of active compound and
excipients into a hard gelatin capsule. Alternatively, a semi-solid matrix of active
compound and high molecular weight polyethylene glycol can be prepared and filled
into a hard gelatin capsule; or a solution of active compound in polyethylene glycol or
a suspension in edible oil, for example liquid paraffin or fractionated coconut oil can
be prepared and filled into a soft gelatin capsule. Compound of Formula I and their
pharmaceutically acceptable salts which are active when given parenterally can be
formulated for intramuscular or intravenous administration.
-
A typical composition for intra-muscular administration will consist of a
suspension or solution of active ingredient in an oil, for example arachis oil or sesame
oil. A typical composition for intravenous administration will consist of a sterile
isotonic aqueous solution containing, for example active ingredient, dextrose, sodium
chloride, a co-solvent, for example polyethylene glycol and, optionally, a chelating
agent, for example ethylenediamine tetracetic acid and an anti-oxidant, for example,
sodium metabisulphite. Alternatively, the solution can be freeze dried and then
reconstituted with a suitable solvent just prior to administration.
-
Compounds of Formula I and their pharmaceutically acceptable salts which are
active on rectal administration can be formulated as suppositories. A typical
suppository formulation will generally consist of active ingredient with a binding
and/or lubricating agent such as a gelatin or cocoa butter or other low melting
vegetable or synthetic wax or fat.
-
Compounds of Formula I and their pharmaceutically acceptable salts which are
active on topical administration can be formulated as transdermal compositions. Such
compositions include, for example, a backing, active compound reservoir, a control
membrane, liner and contact adhesive.
-
The typical daily dose of a compound of Formula I varies according to
individual needs, the condition to be treated and with the route of administration.
Suitable doses are in the general range of from 0.001 to 10 mg/kg bodyweight of the
recipient per day.
-
Within this general dosage range, doses can be chosen at which the
compounds of Formula I lower plasma cholesterol levels and raise metabolic rate with
little or no direct effect on the heart. In general, but not exclusively, such doses will
be in the range of from 0.5 to 10 mg/kg.
-
In addition, within the general dose range, doses can be chosen at which the
compounds of Formula I lower plasma cholesterol levels and have little or no effect
on the heart without raising metabolic rate. In general, but not exclusively, such doses
will be in the range of from 0.001 to 0.5 mg/kg.
-
It is to be understood that the 2 sub ranges noted above are not mutually
exclusive and that the particular activity encountered at a particular dose will depend
on the nature of the compound of Formula I used.
-
Preferably, the compound of Formula I is in unit dosage form, for example, a
tablet or a capsule so that the patient may self-administer a single dose. In general,
unit doses contain in the range of from 0.05-100 mg of a compound of Formula I.
Preferred unit doses contain from 0.05 to 10 mg of a compound of Formula I.
-
The active ingredient may be administered from 1 to 6 times a day. Thus daily
doses are in general in the range of from 0.05 to 600 mg per day. Preferably, daily
doses are in the range of from 0.05 to 100 mg per day. Most preferably from 0.05 to 5
mg per day.
-
As mentioned above, the present invention relates to compounds of Formula I:
where n is 1,2 or 3; R
1 is C
112alkyl, C
3-12alkanol, C
2-6alkenyl, C
5-12alkenol, heterocyclo,
aryl substituted with at least one electron-donating group, -OR
2 or SR
2, where R
2 is
C
1-12alkyl or aryl, or AC(O)NR
12R
13, where A is C
215alkyl or C
415alkenyl and R
12 and
R
13 are C
16alkyl; R
3 and R
5 are methyl; R
4 is hydrogen, C
16alkyl or cycloalkyl; R
6 and
R
9 are hydrogen or C
16alkyl; R
7 and R
8 are independently hydrogen, halogen, C
16alkyl,
optionally substituted phenyl, optionally substituted benzyl, or heteroaryl; with the
proviso that R
7 and R
8 cannot both be hydrogen; R
10 is hydrogen, C
16alkyl, cycloalkyl,
or acyl; and R
11 is hydrogen, C
16alkyl, or cydoalkyl.
-
In preferred embodiments, n is 1. Preferred R4 R6 R7, R9, R10 and R11
substituents are hydrogen. R8 is preferably C16alkyl, for example, isopropyl.
-
The R1 substituent is preferably C2-6alkenyl; phenyl substituted with at least
one, preferably two, electron-donating groups; -OR2 or SR2, where R2 is C1-6alkyl or
phenyl; C3-12alkanol; or AC(O)NR12R13, where A is C215alkyl or C415alkenyl and R12
and R13 are C16alkyl. A preferred -OR2 substituent is ethoxy. Preferred SR2
substituents include ethylthio and phenylthio.
-
Another preferred R1 substituent is C2-6alkenyl, for example -CH2-CH=CH2. A
preferred R1 substituent is also a phenyl substituted with at least one, preferably two,
electron-donating groups such as methoxy, for example, dimethoxyphenyl. A
preferred C3-12alkanol is -(CH2)3-OH. Preferred AC(O)NR12R13 groups include
-(CH2)10-C(O)-N(CH3)-(CH2)3(CH3) and
(CH2)2-C=C-(CH2)6-C(O)-N(CH3)-(CH2)3(CH3)-
METHODS OF PREPARATION
-
Compounds of Formula I are prepared from an intermediate of Formula II or
Formula IIa, which is formed from the intermediates (3) and (6), the preparation of
which is shown below. Suitable protecting groups for the X substituent of Formula II
include, but are not limited to, silyl containing protecting groups such as TIPS.
Suitable protecting groups for the Y substituent of Formula II include, but are not
limited to, -OT where T is C1-6alkyl such as methyl, and O(CH2)nCO2Me and
O(CH2)nCO2Et.
1. Preparation of Compounds of Formula II
1a. Preparation of Compounds of Formula (3)
-
Compounds of Formula (3) are prepared as shown below in Reaction Scheme
-
Compounds of Formula (1) are commercially available, or may be prepared by
means well known in the art. In general, the phenol of Formula (1) is first protected
by conversion to the methoxy derivative, for example by reacting (1) with methyl
iodide in the presence of a base, for example potassium carbonate, in a polar solvent,
for example DMF. When the reaction is substantially complete, the protected phenol
of Formula (2) is isolated and purified by conventional means, preferably by flash
chromatography.
-
Clearly, other conventional phenol protecting groups could be utilized instead
of methoxy, for example a silyl protecting group, e.g. t-butyldimethylsilyloxy.
-
The compound of Formula (2) is then brominated using potassium bromide in
the presence of a crown ether, for example 18-crown-6, and an oxidizing agent, for
example 3-chloroperoxy benzoic acid. The reaction is carried out in an inert solvent,
preferably CH2Cl2. When the reaction is substantially complete, the 4-bromo
derivative of Formula (3) is isolated and purified by conventional means, preferably
by flash chromatography.
1b. Preparation of Compounds of Formula (6)
-
Compounds of Formula (6) are prepared as shown below in Reaction Scheme
II.
-
Compounds of Formula (4) are commercially available, or may be prepared by
means well known in the art. In general, the phenol of Formula (4) is first protected
by conversion to the methoxy derivative, or other conventional phenol protecting
groups, as disclosed in Reaction Scheme I above, to give a p-bromo compound of
Formula (5).
-
The bromo moiety of the compound of Formula (5) is then converted to a
formyl group. The reaction is carried out conventionally, adding t-butyllithium to a
solution of (5) in an inert solvent at about -78°C, preferably THF, and adding DMF to
the cold solution. After stirring cold, the mixture is allowed to warm to room
temperature. When the reaction is substantially complete, the 4-formyl derivative of
Formula (6) is isolated and purified by conventional means, preferably by flash
chromatography.
1c. Preparation of Compounds of Formula II from Compounds of Formulas (3) and
(6)
-
Compounds of Formula II are prepared from (3) and (6) as shown below in
Reaction Scheme III.
-
Compounds of Formula (7) are prepared by reaction of (3) and (6). In general,
the p-bromo compound of Formula (3) is dissolved in an inert solvent, preferably
THF, cooled to about -78°C, and t-butyllithium added. After stirring for about 10
minutes, the compound of Formula (6) is added. After stirring cold, the mixture is
allowed to warm to room temperature. When the reaction is substantially complete,
the carbinol derivative of Formula (7) is isolated and purified by conventional means,
preferably by flash chromatography.
-
Solvolysis of the benzylhydroxy group of the compound of Formula (7) yields
the compound of Formula II. In general, the reaction is carried out with CH2Cl2 in an
acidic medium, preferably trifluoroacetic acetic acid ("TFA"), under an inert
atmosphere at about -45°C in the presence of a suitable acid-stable nucleophilic
species, as described in Luengo, et al., J. Org. Chem. 59:6512 (1994) and Luengo, et
al., Chem. Biol. 2:471 (1995). Particularly suitable nucleophiles include ethanol,
allyltrimethylsilane, 1,3dimethoxybenzene, ethanethiol and thiophenol. When the
reaction is substantially complete, the compound of Formula II is isolated by
conventional means, and preferably used with no further purification.
2. Preparation of Compounds of Formula I from Compounds of Formula II
-
Compounds of Formula I are prepared from II as shown below in Reaction
Scheme IV.
-
The dimethoxy derivative of Formula II is demethylated. The reaction is
carried out conventionally, using boron tribromide in CH2Cl2. When the reaction is
substantially complete, the dihydroxy derivative of Formula (8) is isolated and
purified by conventional means, preferably by flash chromatography.
-
The compound of Formula (8) is then converted to a compound of Formula I
where R10 is hydrogen by reaction with an ester of formula Q-(CH2)n-CO2R11, where Q
is chloro, bromo or iodo, n is 1, 2 or 3, and R11 is lower alkyl, for example t-butyl.
The compound of Formula (8) is dissolved in an inert solvent, for example THF,
cooled to about 25°C, and cesium carbonate (Cs2CO3) added followed by the halo
ester. The mixture is stirred cold for about 1 hour, then allowed to warm to room
temperature. When the reaction is substantially complete, the ester derivative of a
compound of Formula I is isolated and purified by conventional means, preferably by
flash chromatography. This ester is dissolved in a protic solvent, preferably methanol,
and hydrolysed with a base, preferably sodium hydroxide. After acidification, the
compound of Formula I is isolated and purified by conventional means.
-
In addition, compounds of Formula I can also be prepared from an
intermediate of Formula III, which is formed from intermediates of Formula (10) and
(13), the preparation of which is shown below.
3. Preparation of Compounds of Formula III
-
Suitable protecting groups for the X substituent of Formula III include, but are
not limited to, silyl containing protecting groups such as TIPS. Suitable protecting
groups for the Y substituent of Formula III include, but are not limited to,
O(CH2)nCO2Me, O(CH2)nCO2Et and OT' where T' is a silyl containing protecting
group such as TBMPS. Suitable Z' groups in Formula III include, but are not limited
to hydroxy and lower alkoxy, for example, methoxy and ethoxy.
-
This synthesis includes introduction of the TIPS and TBMPS protecting
groups to the phenolic hydroxyls. For purposes of illustration, the following reaction
schemes illustrate the synthesis of a compound of Formula I where n is 1, R8 is
isopropyl and R10 and R11 are hydrogen. It is understood that by replacing the starting
materials with other compounds of Formulas (10) and (13), and following the
procedures described below, other compounds of Formula I are prepared.
3a. Preparation of Compounds of Formula (10)
-
Compounds of Formula (10) are prepared as shown below in Reaction Scheme
V.
-
Compounds of Formula (9) are readily synthesized by the addition of 1
equivalent bromine in CH2Cl2 to the commercially available 2-isopropyl phenol. In
general, the phenol of Formula (9) is first protected by conversion to the
triisopropylsilyloxy derivative with TIPS chloride and imidazole and ClCH2CH2Cl, to
give a p-bromo compound of Formula (10). When the reaction is substantially
complete, the p-bromo compound is isolated and purified by conventional means,
preferably by flash chromatography.
3b. Preparation of Compounds of Formula (13)
-
Compounds of Formula (13) are prepared as shown below in Reaction Scheme
VI.
-
Compounds of Formula (11) are commercially available, or may be prepared
by means well known in the art. In general, the phenol of Formula (11) is first
protected by conversion to the tert-butylmethoxyphenylsilyloxy derivative with
TBMPS bromide in imidazole and CH2Cl2, to give a p-bromo compound of Formula
(12).
-
The bromo moiety of the compound of Formula (12) is then converted to a
formyl group. The reaction is carried out conventionally, adding n-butyllithium to a
cold solution of (12) in an inert solvent (about -78°C), preferably THF, and adding
DMF to the cold solution. After stirring cold, the mixture is allowed to warm to room
temperature. When the reaction is substantially complete, H3O+ is added. The
4-formyl derivative of Formula (13) is then isolated and purified by conventional
means, preferably by flash chromatography.
3c. Preparation of Compounds of Formula III from Compounds of Formulas (10) and
(13)
-
Compounds of Formula III are prepared from (10) and (13) as shown below in
reaction Scheme VII.
-
In general, the p-bromo compound of Formula (10) and 1 eq. n-butyllithium
are mixed together in a cold inert solvent (e.g., THF at about -78°C). After stirring for
about 10 minutes, this mixture is then added to a suspension of CeCl3, in a cold inert
solvent (e.g., THF at about 78°C) and stirred for about 30 minutes. Then the
compound of Formula (13) is added. The reaction is allowed to continue in cold THF
(about -78°C). After stirring cold, the mixture is allowed to warm to room
temperature. When the reaction is substantially complete, H3O+ is added. The
compound of Formula (14) is isolated and purified by conventional means, preferably
by flash chromatography.
-
Solvolysis of the benzylhydroxy group of the compound of Formula (14)
yields the compound of Formula III. In general, the reaction is carried out with
CH2Cl2 in an acidic medium, preferably TFA, under an inert atmosphere at about
-45°C in the presence of a suitable acid-stable nucleophilic species, as described in
Luengo, et al., J. Org. Chem. 59:6512 (1994) and Luengo, et al., Chem. Biol. 2:471
(1995). Particularly suitable nucleophiles include ethanol, methanol,
allyltrimethylsilane, 1,3-dimethoxybenzene, ethanethiol and thiophenol. When the
reaction is substantially complete, the compound of Formula III is isolated and
purified by conventional means, preferably by flash chromatography.
4. Preparation of Compounds of Formula I from Compounds of Formula III
-
Compounds of Formula I are prepared as shown below in Reaction Scheme
VIII.
-
The TBMPS group is removed from the compound of Formula III under
conditions that do not harm the sensitive heteroatomic bridge substituents. TBMPS is
selectively cleaved with stoichiometric Et3N(3HF in the presence of an inert solvent
such as THF (to yield Z' = -OMe as shown in Scheme VII) or stoichiometric
tetrabutylammonium fluoride (1 eq. TBAF) in CH2Cl2 to yield Z' = -OEt), at about
-78°C. The mixture is washed with a saturated ammonium chloride solution and the
aqueous phase extracted with Et2O to yield the phenol of Formula (15).
-
The resulting phenol is alkylated with an ester of formula Q-(CH2)n-CO2R11,
where Q is chloro, bromo or iodo, n is 1, 2 or 3, and R11 is lower alkyl, for example
ethylbromoacetate or methylbromoacetate, Cesium carbonate and the halo ester are
added to the phenol in a DMF solution. The mixture is stirred cold for about 1 hour
then mixed with a saturated ammonium chloride solution and extracted with Et2O to
yield the ester of Formula III (Compound 16), which is then isolated by conventional
means.
-
The next reaction is carried out with CH2Cl2 in an acidic medium, preferably
TFA at about -45°C, by the addition of a compound of formula R1-H to the compound
of Formula (15). R1 is a lower alkenyl group, an optionally substituted phenyl group,
an -OR2 or a -SR2 group, where R2 is lower alkyl or phenyl. If the final product has
R1= -OCH2CH3, then this step is optional since the desired R1 substitution may be
made during the synthesis of the compound of Formula III.
-
The ester of Formula (16) is then dissolved in a protic solvent, preferably
methanol, and hydrolysed with a base, preferably lithium hydroxide. The mixture is
washed with a saturated ammonium chloride solution and the aqueous phase extracted
with Et2O to yield the phenol of Formula (17). Subsequent deprotection of the phenol
is achieved by dissolving the phenol in equal parts of acetonitrile and CH2Cl2,
followed by the addition of 1 eq. potassium fluoride in 0.5 eq. 18-crown-6, to yield the
compound of Formula I, which is isolated and purified by conventional means such as
reverse phase HPLC.
5. Preparation of Compounds of Formula I, where R
1
is A-C(Q)NR
12
R
13
and A is
alkyl (I-1)
-
Compound (I-1) and similar compounds are prepared as shown below in
Reaction Scheme IX.
6. Preparation of Compounds of Formula I, where R 1 is -A-C(O)NR 12 R 13 and A is alkenyl (I-2)
-
Compound (I-2) and similar compounds are prepared as shown below in
Reaction Scheme X
7. Preparation of Compounds of Formula I, where R
1
is a lower alkenyl (I-3)
-
Compound (I-3) and similar compounds are prepared as shown below in
Reaction Scheme XI
8. Preparation of Compounds of Formula I, where R
1
is a lower alkoxy (I-4)
-
Compound (I-4) and similar compounds are prepared as shown below in
Reaction Scheme XII
9. Preparation of Compounds of Formula I where R 1 is a substituted aryl (I-5)
-
Compound (I-5) and similar compounds are prepared as shown below in
Reaction Scheme XIII.
Isolation and Purification of the Compounds
-
Isolation and purification of the compounds and intermediates described
herein can be effected, if desired, by any suitable separation or purification procedure
such as, for example, filtration, extraction, crystallization, colunm chromatography,
thinlayer chromatography, thicklayer chromatography, preparative low or
highpressure liquid chromatography or a combination of these procedures. Specific
illustrations of suitable separation and isolation procedures can be had by reference to
the Examples hereinbelow. However, other equivalent separation or isolation
procedures could, of course, also be used.
Separation of Enantiomers
-
The enantiomers of the compounds and intermediates described herein can be
effected, if desired, by any conventional resolution means, for example by separation
(e.g. fractional crystallization) of the diastereomeric salts formed by the reaction of a
racemic compound of Formula I with an optically active base.
Salts of Compounds of Formula I
-
The compounds of Formula I where R11 is hydrogen may be converted to a
corresponding base addition salt from inorganic and organic bases by conventional
means. Typically, the free acid of Formula I is dissolved in an inert organic solvent
such as diethyl ether, ethyl acetate, chloroform, ethanol or methanol and the like, and
the base added in a similar solvent. The temperature is maintained at 050°C. The
resulting salt precipitates spontaneously or may be brought out of solution with a less
polar solvent.
-
The following preparation and examples illustrate the invention but are not
intended to limit its scope.
General Methods
-
Proton and carbon-13 nuclear magnetic resonance spectra (1H NMR,13C
NMR) were obtained on a General Electric QE-300 (300 Mhz) spectrometer, with
tetramethylsilane used as the reference.
-
Flash chromatography on crude products was performed using 230-400 mesh
silica gel (Aldrich Chemical Co.). Purity of compounds was determined by TLC
using commercial silica gel plates (Alltech, Alugram® Sil G/UV 254) and by 1H
NMR and HRMS.
-
Methylene chloride (anhydrous) (CH2Cl2), THF (anhydrous) and reagents were
purchased from Aldrich Chemical Co. and used without further purification. Unless
specified otherwise, reactions were performed under Argon inert atmosphere.
Abbreviations
-
- DMF
- N,N-dimethylformamide
- HBTU
- O-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyl-uronium
hexafluorophosphate
- TBAF
- tetrabutylammonium fluoride
- TBMPS
- tert-butylmethoxyphenylsilyloxy
- TEMPO
- 2,2,6,6-tetramethylpiperidine-N-oxide radical
- TFA
- trifluoroacetic acetic acid
- THF
- tetrahydrofuran
- TIPS
- triisopropylsilyl
EXAMPLE 1
Preparation of Compounds of Formula (2)
Preparation of (2) where R
7
is Isopropyl; and R
6
, R
8
, R
9
are H
-
A mixture of 2-isopropylphenol (a compound of Formula 1) (12.0 g, 88.1
mmol), methyl iodide (25.0 g, 176.2 mmol), and potassium carbonate (24.3 g, 176.2
mmol) in 44 mL of DMF was stirred for 20 hours at room temperature. The reaction
mixture was diluted with 300 mL of ether and washed with 250 mL of water and
5x100 mL of brine. The organic portion was dried (MgSO4), filtered, and evaporated
to give an oil, which was purified by flash column chromatography (silica gel, 90:10
hexane/ethylacetate) to give 2isopropylanisole (a compound of Formula 2) (12.5 g,
82.1 mmol, 93%); 1HNMR (CDCl3) (1.2 (d, 6H), 3.3 (heptet, 1H), 3.8 (s, 3H), 6.8 (d,
1H), 6.88 (t, 1H), 7.13 (d, 1 H), 7.2 (t, 1H).
Preparation of (2), varying R
6
, R
7
, R
8
, R
9
-
In a similar manner, replacing 2-isopropylphenol with other compounds of
Formula (1) and following the procedure described in Example 1 above, other
compounds of Formula (2) are prepared.
EXAMPLE 2
Preparation of Compounds of Formula (3)
Preparation of (3) where R
7
is Isopropyl; and R
6
, R
8
, R
9
are H
-
To a suspension of potassium bromide (18.8 g, 157.7 mmol) in 400 mL of
CH2Cl2 at 0°C were added 18-Crown-6 (2.08 g, 7.88 mmol), 3-chloroperoxy benzoic
acid (27.2 g, 157.7 mmol) and 2isopropylanisole (a compound of Formula 1 from
Example 1) (12.0 g, 78.8 mmol). After stirring for 3 hours at 0°C, the reaction mixture
was poured into ice water (500 mL), and stirred for 30 minutes. The organic layer was
separated, washed with saturated NaHCO3 solution (400 mL), followed by water (300
mL), and dried (MgSO4). The solvent was evaporated to give an oil, which was
purified by flash column chromatography (silica gel, 98:2 hexane/ethylacetate) to give
13 g (56.7 mmol, 72%) of 4-bromo-2isopropylanisole (a compound of Formula 3) as
an oil; 1HNMR (CDCl3) (1.2 (d, 6H), 3.3 (heptet, 1H), 6.7 (d, 1H), 6.84 (d, 1H), 7.29
(s, 1H).
Preparation of (3), varying R
6
, R
7
, R
8
, R
9
-
In a similar manner, replacing 2-isopropylanisole with other compounds of
Formula (2) and following the procedure described in Example 2 above, other
compounds of Formula (3) are prepared.
EXAMPLE 3
Preparation of Compounds of Formula (5)
Preparation of (5) where R
4
is H; and R
3
R
5
are Methyl
-
A mixture of commercially available 4-bromo-3,5-dimethylphenol (a
compound of Formula 4) (25.0 g, 124.3 mmol), methyl iodide (35.3 g, 248.6 mmol),
and potassium carbonate (34.4 g, 248.6 mmol) in 62.5 mL of DMF was stirred for 2
hours at room temperature. The reaction mixture was diluted with 300 mL of ether
and washed with 250 mL of water and 5x100 mL of brine. The organic portion was
dried (MgSO4) filtered, and evaporated to give an oil, which was purified by flash
column chromatography (silica gel, 90:10 hexane/ethylacetate) to give
4-bromo-3,5-dimethylanisole (a compound of Formula 5) (26 g, 120.8 mmol, 97%);
1HNMR (CDCl3) (2.39 (s, 6H), 3.76 (s, 3H), 6.67 (s, 2H).
Preparation of (5), varying R
3
, R
4
R
5
-
In a similar manner, replacing 4-bromo-3,5-dimethylphenol with other
compounds of Formula (4) and following the procedure described in Example 3
above, other compounds of Formula (5) are prepared.
EXAMPLE 4
Preparation of Compounds of Formula (6)
Preparation of (6) where R
4
is H; and R
3
, R
5
are Methyl
-
To 4-bromo-3,5-dimethylanisole (a compound of Formula 5 from Example 3)
(20 g, 93.0 mmol) in 500 mL of THF at -78°C was added 120 mL of tert-Butyllithium
(1.7 M in pentane). The reaction mixture was stirred for 30 minutes at -78°C and then
DMF (136.0 g, 186.0 mmol) was added. The reaction mixture was stirred for 1 hour at
-78°C and for 1.5 hours at room temperature, diluted with 300 mL of ether, washed
with 300 mL of water, acidified 1N HCl, and 5x100 mL of brine. The organic portion
was dried (MgSO4) filtered, and evaporated to give the crude product, which was
purified by flash column chromatography (silica gel, 90:10 hexanelethylacetate) to
yield 2,6-dimethyl-4-methoxybenzaldehyde (a compound of Formula 6), (9.50 g, 57.8
mmol, 62%) as a white solid; 1HNMR (CDCl3 (2.61 (s, 6H), 3.83 (s, 3H), 6.6 (s, 2H),
10.5 (s, 1H).
Preparation of (6), varying R
5
-
In a similar manner, replacing 4-bromo-3,5-dimethylanisole with other
compounds of Formula (5) and following the procedure described in Example 4
above, other compounds of Formula (6) are prepared.
EXAMPLE 5
Preparation of Compounds of Formula (7)
Preparation of (7) where R
4
, R
6
R
7
, R
9
are H; R
3
, R
5
are Methyl; and R
8
is Isopropyl
-
To 4-bromo-2isopropylanisole (a compound of Formula 3 from Example 2)
(12 g, 52.4 mmol) in 300 mL of THF at -78°C was added 68 mL of tert-butyllithium
(1.7 M in pentane). The reaction mixture was stirred for 10 min at -78°C and then
2,6-dimethyl-4-methoxybenzaldehyde (a compound of Formula 6 from Example 4)
(8.6 g, 52.4 mmol) was added. The reaction mixture was stirred for 1 hour at -78°C
and for 1.5 hours at room temperature, diluted with 150 mL of ether, washed with 150
mL of water, acidified with 1N HCl, and washed with 5x50 mL of brine. The organic
portion was dried (MgSO4), filtered, and evaporated to give the crude product, which
was purified by flash column chromatography (silica gel, 95:5 hexane/ethylacetate) to
yield 3,5-dimethyl-4-(3'-isopropyl-4'-methoxybenzylhydroxy) anisole (7) (12 g, 38.2
mmol, 73%) as an oil; 1HNMR (CDCl3) (1.2 (dd, 6H), 2.27 (s, 6H), 3.30 (heptet, 1H),
3.80 (s, 6H), 6.26 (s, 1 H), 6.59 (s, 2H), 6.76 (d, 1H), 6.89 (d, 1H), 7.24 (s, 1H).
Preparation of (7), varying R
4
, R
6
, R
7
, R
8
, R
9
-
In a similar manner, optionally replacing 4-bromo-2isopropylanisole with
other compounds of Formula (3), and optionally replacing
2,6-dimethyl-4-methoxybenzaldehyde with other compounds of Formula (6), and
following the procedure described in Example 5 above, other compounds of Formula
(7) are prepared.
EXAMPLE 6
Preparation of Compounds of Formula II
Preparation of II where R
4
, R
6
, R
7
, R
9
are H; R
3
, R
5
are Methyl; R
8
is Isopropyl; and R
1
is Ethoxy
-
A solution of 3,5-dimethyl-4-(3'-isopropyl-4'-methoxybenzylhydroxy) anisole
(a compound of Formula 7 from Example 5) (40 mg, 0.13 mmol) and ethanol (5.1
mmol) in CH2Cl2 (8 mL) was cooled to -45 °C (dry ice/acetonitrile bath). TFA (167
(L, 2.2 mmol) was added and the reaction stirred 2 h at -45 °C. The reaction was
quenched by adding sat. NaHCO3 (5 mL) and water (5 mL). Layers were separated
and the aqueous phase extracted twice with diethyl ether ("Et2O") (7 mL). Combined
extracts were washed with brine (10ml), dried over MgSO4 and evaporated to give the
crude product, which was purified by flash chromatography (1:20 Et2O-hexanes) to
yield ethoxy-4,4'-dimethoxy-2,6-dimethyl-3'-(1- methylethyl)diphenylmethane (a
compound of Formula II) (40 mg, 0.095mmol, 73%); 1H NMR (CDCl3) (7.17 (d, J=
1.6 Hz, 1 H), 6.90 (dd, J= 1.5, 8.4 Hz, 1 H), 6.71 (d, J= 8.4Hz, 1 H),6.57 (s, 2 H),
5.80 (s, 1 H), 3.79 (s, 3 H), 3.78 (s, 3 H), 3.47 (br q, J= 7.0 Hz, 2 H),3.26 (heptet, J=
6.9 Hz, 1 H), 2.24 (s, 6 H), 1.26 (t, J= 7.0 Hz, 3 H), 1.17 (d, J= 6.7 Hz, 3 H), 1.15 (d,
J = 6.7 Hz, 3 H); HRMS exact mass calcd for C22H30O3 342.2195, found 342.2189.
Preparation of II, varying R
1
R
1
is ethylthio
-
Substitution of ethanethiol for ethanol in the above reaction and starting with
35 mg of the compound of Formula (7), with the additional step of quenching the
reaction with 0.5 M NaOH (10 mL), and treating the extracted aqueous phase with
bleach to reduce the stench, yielded
ethylthio-4,4'-dimethoxy-2,6-dimethyl-3'-(1-methylethyl)diphenylmethane (a
compound of Formula II) (33 mg, 0.098 mmol, 89%); 1H NMR (CDCl3) (7.30 (d, J=
2.0 Hz, 1 H), 7.05 (dd, J= 1.6, 8.4 Hz, 1 H), 6.73 (d, J= 8.5 Hz, 1 H), 6.56 (s, 2 H),
5.58 (s, 1 H), 3.79 (s, 3 H), 3.77 (s, 3 H), 3.27 (heptet, J= 6.9 Hz, 1 H), 2.67-2.48 (M,
2 H), 2.23 (br s, 6 H), 1.28 (t, J= 7.4 Hz, 3 H), 1.17 (app t, J= 7.0 Hz, 6 H); HRMS
exact mass calcd for C22H30O2S 358.1966, found 358.1953.
R
1
is phenylthio
-
Substitution of thiophenol for ethanol in the above reaction and starting with
41 mg (0.13 mmol) of the compound of Formula (7), with the additional step of
quenching the reaction with 0.5 M NaOH (10 mL), and treating the extracted aqueous
phase with bleach to reduce the stench, yielded
4,4'-dimethoxy-2,6-dimethyl-3'-(1-methylethyl)diphenylphenylthio methane (a
compound of Formula II) (35 mg, 0.073 mmol, 66%); 1H NMR (CDCl3) (7.34 (s, 1
H), 7.32 (s, 1 H), 7.25-7.17 (m, 3 H), 7.08 (dd, J= 1.7, 8.4 Hz, 1 H), 6.72 (d, J= 8.5
Hz, 1 H), 6.55 (s, 2 H), 5.89 (s, 1 H), 3.79 (s, 3 H), 3.78 (s, 3 H), 3.26 (heptet, J= 6.9
Hz, 1 H), 2.13 (br s, 6 H), 1.15 (d, J= 6.9 Hz, 3 H), 1.09 (d, J= 6.9 Hz, 3 H); HRMS
exact mass calcd for C26H29O2S (M - H+) 405.1888, found 405.1894.
R
1
is alkenyl
-
Substitution of allyltrimethylsilane (CH2CHCH2Si(CH3)3) for ethanol in the
above reaction and starting with 35 mg of the compound of Formula (7), yielded
4,4-[4',4"-dimethoxy-2',6'-dimethyl-3'-(1-methylethyl)diphenyl]butan-1-ene (a
compound of Formula II) (41 mg, 0.12 mmol, 93%); 1H NMR (CDCl3) (7.05 (d, J=
1.7 Hz, 1 H), 6.87 (dd, J = 1.6, 8.4 Hz, 1 H), 6.71 (d, J= 8.5 Hz, 1 H), 6.54 (s, 2 H),
5.78-5.67 (m, 1 H), 5.09 (dd, J= 1.0, 17.1 Hz, 1 H), 4.93 (d, J = 1 0.2 Hz, 1 H), 4.50
(t, J = 7.9 Hz, 1 H), 3.78 (s, 3 H), 3.77 (s, 3 H), 3.26 (heptet, J = 6.9 Hz, 1 H),
3.09-3.00 (m, 1 H), 2.80-2.70 (m, 1H), 2.15 (br s, 6 H), 1.16 (d, J = 7.1 Hz, 3 H), 1.14
(d, J = 7.1 Hz, 3H); HRMS exact mass calcd for C23H30O2 338.2246, found 338.2247.
R
1
is a substituted aryl such as 2.4-dimethoxyphenyl
-
Substitution of 1,3-dimethoxybenzene for ethanol in the above reaction,
yielded 4,4',2",4"-tetramethoxy-2,6-dimethyl-3'-(1-methylethyl)triphenylmethane (a
compound of Formula II) (90% yield); 1H NMR (CDCl3) (6.85 (s, 1 H), 6.76 (d, J=
8.4 Hz, 1 H), 6.68 (s, 1 H), 6.54 (s, 2 H), 6.47 (d, J = 2.2 Hz, 1H), 6.36 (dd, J = 2.3,
8.5 Hz, 1 H), 5.93 (s, 1 H), 3.79 (s, 3 H), 3.78 (s, 3 H), 3.77 (s, 3 H), 3.67 (s, 3 H),
3.24 (heptet, J= 6.9 Hz, 1 H), 1.99 (s, 6 H), 1.09 (d, J= 6.9 Hz, 6 H); HRMS exact
mass calcd for C28H34O4 434.2457, found 434.2458.
Preparation of II, varying R
4
, R
6
, R
7
, R
8
, R
9
-
In a similar manner, replacing
3,5-dimethyl-4-(3'-isopropyl-4'-methoxybenzylhydroxy) anisole with other
compounds of Formula (7), and following the procedure described in Example 6
above, other compounds of Formula II are prepared.
EXAMPLE 7
Preparation of Compounds of Formula (15)
Preparation of (15) where R
4
, R
6
, R
7
, R
9
are H; R
3
R
5
are Methyl; R
8
is Isopropyl; R
1
is Ethoxy
-
To
ethoxy-4-[(1,1-dimethylethyl)methoxyphenylsilyloxy]-2,6-dimethyl-4'-tris(1-methylet
hyl)silyloxy-3'-(1-methylethyl)diphenylmethane (a compound of Formula III) is added
anhydrous CH2Cl2. The reaction mixture is cooled to -78°C and 1.0 eq. TBAF is
added via syringe. The reaction is allowed to warm slowly and is stirred. The
reaction mixture is then washed with saturated ammonium chloride and the aqueous
phase extracted with Et2O. The combined organic layers are washed with brine, dried
(MgSO4), and evaporated to give the crude product. Purification by flash
chromatography (silica gel, 1:9 ethyl acetate:hexanes) gives
ethoxy-4-hydroxy-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethyl)diphe
nylmethane (a compound of Formula 15).
Preparation of (15), varying R
1
, R
4
, R
6
, R
7
, R
8
, R
9
-
In a similar manner, replacing
ethoxy-4-[(1,1-dimethylethyl)methoxyphenylsilyloxy]-2,6-dimethyl-4'-tris(1-methylet
hyl)silyloxy-3'-(1-methylethyl)diphenylmethane with other compounds of Formula III,
and following the procedure described in Example 7 above, other compounds of
Formula (15) are prepared.
EXAMPLE 8
Preparation of Compounds of Formula (16)
Preparation of (16) where R 4 R 6 R 7 , R 9 are H; R 3 R 5 are Methyl; R 8 is Isopropyl: R 1 is Ethoxy
-
To
ethoxy-4-hydroxy-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethyl)diphe
nylmethane (a compound of Formula 15 from Example 7), is added DMF, 2 eq.
Cs2CO3 and 1.5 eq. ethylbromoacetate. The reaction is stirred, then mixed with a
saturated ammonium chloride solution and extracted with Et2O. The combined
organics are dried (MgSO4 and evaporated to give the crude product,
ethoxy-4-ethyloxyacetate-2,6-dimethyl-4'-tris(1-methylethyl) silyloxy-3'-(1-methylethy
1)diphenylmethane (a compound of Formula 16).
Preparation of (16), varying R
1
, R
4
, R
6
, R
7
, R
8
, R
9
-
In a similar manner, replacing
ethoxy-4-hydroxy-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethyl)diphe
nylmethane with other compounds of Formula (15), and following the procedure
described in Example 8 above, other compounds of Formula (16) are prepared.
EXAMPLE 9
Preparation of Compounds of Formula (17)
Preparation of (17) where R
4
, R
6
, R
7
, R
9
are H; R
3
, R
5
are Methyl, R
8
is Isopropyl; R
1
is Ethoxy
-
To
ethoxy-4-ethyloxyacetate-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethy
1)diphenylmethane (a compound of Formula 16 from Example 8) is added methanol, 2
eq. lithium hydroxide monohydrate and 1 eq. water. The reaction is stirred, then the
solvent volume is reduced by evaporation. The residue is suspended in a saturated
ammonium chloride solution and extracted with Et2O. The combined organics are
dried (MgSO4) and evaporated to give the crude product,
ethoxy-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethyl)diphenylmethane
-4-oxyacetic acid (a compound of Formula 17).
Preparation of (17), varying R
1
, R
4
, R
6
, R
7
, R
8
, R
9
-
In a similar manner, replacing
ethoxy-4-ethyloxyacetate-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethy
1)diphenylmethane with other compounds of Formula (16), and following the
procedure described in Example 9 above, other compounds of Formula (17) are
prepared.
EXAMPLE 10
Preparation of Compounds of Formula I
Preparation of I where n is 1; R
4
, R
6
, R
7
, R
9
, R
10
are H; R
3
, R
5
are Methyl: R
8
is
Isopropyl; R
1
is Ethoxy
-
Ethoxy-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethyl)diphenyl
methane-4-oxyacetic acid (a compound of Formula 17 from Example 9) is dissolved
in an equal mixture of acetonitrile and CH2CH2. 18-Crown-6 (0.5 eq.) and potassium
fluoride (1.1 eq.) are added and the reaction mixture is stirred for 15 hours. The
solvent volume is reduced by evaporation and the residue is suspended in a saturated
ammonium chloride solution and extracted with CHCl3. The combined organics are
dried (MgSO4) and evaporated to give the crude product. Reverse phase (C18) HPLC
(water/acetonitrile + 0.1 % TFA) gives the pure product,
ethoxy-2,6-dimethyl-4'-hydroxy-3'-(1-methylethyl)diphenylmethane-4-oxyacetic acid
(a compound of Formula I).
Preparation of I, varying n, R
1
, R
4
, R
6
, R
7
, R
8
, R
9
, R
10
-
In a similar manner, replacing
ethoxy-2,6-dimethyl-4'-tris(1-methylethyl)silyloxy-3'-(1-methylethyl)diphenylmethane
-4-oxyacetic acid with other compounds of Formula (17), and following the procedure
described in Example 10 above, other compounds of Formula I are prepared.
EXAMPLE 11
Preparation of Compound (I-1), a Compound of Formula I
Preparation of a Compound of Formula (9)
-
To a stirred, chilled (0(C) solution of commercially available 2-isopropyl
phenol (30.0g, 220mmol) in anhydrous methylene chloride (250ml), elemental
bromine (10ml, 19mmol) was added. After stirring for 1 hr. at 0(C, the reaction was
quenched with NH4O (50ml), water (250ml) and saturated NaHCO3 (200ml). The
aqueous phase was extracted with diethyl ether and combined organic fractions were
washed with brine and dried (MgSO4) followed by filtration; solvent was removed by
rotary evaporation to yield a golden oil (42g). 1H NMR analysis indicated only ~75%
reaction, thus the crude product was subjected to the same reaction conditions with
only 2.8ml (54mmol) of bromine added, which yielded a golden oil (50.6g).
Purification by column chromatography (4" x 7", 5% (20% EtOAc/Hexanes) yielded
4-bromo-2-isopropylphenol (a compound of Formula 9) as a colorless oil (30g, 63%).
1H NMR (600 MHz, CDCl3) (7.28 (d, J= 2.6 Hz, 1 H), 7.15 (dd, J= 8.4, 2.2 Hz, 1
H), 6.63, (d, J = 8.4 Hz, 1 H), 4.79 (s, 1 H), 3.17 (heptet, J = 6.9 Hz, 1 H), 1.24 (d, J =
7.0 Hz, 6 H).
Preparation of a Compound of Formula (10)
-
To a stirred solution of TIPS chloride (12ml, 56mmol) in anhydrous
1,2-dichloroethane (70ml) was added 4-bromo-2-isopropylphenol (a compound of
Formula 9) (9.6g, 46mmol) and imidazole (7.8g, 114mmol). Reaction was refluxed
from 30min, then allowed to stir overnight at room temperature. Reaction was
refluxed 1 hour further, then 150ml of 0.6M HCl was added, layers separated and the
aqueous phase extracted with diethyl ether. Combined organic fractions were washed
with saturated NaHCO3 dried (MgSO4) filtered through a Celite plug and solvent was
removed by rotary evaporation to yield an oil (18.3g). Purification by fractional
distillation (bp 137(C, 0.3mm) yielded
3-isopropyl-4-triisopropylsilyoxy-bromobenzene (a compound of Formula 10) as a
white solid (12.3g, 72%).1H NMR (600 MHz, CDCl3) (7.27 (d, J= 2.6 Hz, 1 H), 7.11
(dd, J = 8.4, 2.6 Hz, 1 H), 6.64 (d,J= 8.8 Hz, 1 H), 3.33 (heptet, J= 6.8 Hz, 1 H), 1.30
(heptet, J= 7.5 Hz, 3 H), 1.19 (d, J = 6.6 Hz, 6 H), 1.10 (d, J= 7.7 Hz, 18 H).
Preparation of a Compound of Formula (12)
-
To a stirred solution of commercially available 4-bromo-3,5-dimethylphenol (a
compound of Formula 11, 13.5g, 66.9mmol) and imidazole (11.4g, 167mmol) in
anhydrous methylene chloride (125ml) was added tert-butylmethoxyphenylsilyl
bromide (19.2g, 70.3mmol). Reaction was stirred 2.5 hr., then 200ml 0.1M HCl was
added and the layers separated. The aqueous layer was extracted with diethyl ether,
and the combined organic fractions were washed with brine and dried (MgSO4).
Removal of drying agent by filtration and solvent by rotary evaporation yielded
4-TBMPS-2,6-dimethylbromobenzene (a compound of Formula 12), as a golden oil
(26.6g, 100%) which was used without further purification. 1H NMR (600 MHz,
CDCl3) (7.66 (d, J = 7.3 Hz, 2 H), 7.44 (t, J = 8.8 Hz, 1 H), 7.39 (t, J= 7.5 Hz, 2 H),
6.73 (s, 2 H), 3.61 (s, 3 H), 2.33 (s, 6 H), 0.99 (s, 9 H).
Preparation of a Compound of Formula (13)
-
To a solution of 4-TBMPS-2,6-dimethylbromobenzene (a compound of
Formula 12) (26.2g, 66.6mmol) in anhydrous THF cooled to -78(C was added a
solution of butyllithium in hexanes (2.5M, 76.6mmol) over several min. Anhydrous
DMF (7.7ml, 99.9mmol) was then added over 5 min. After stirring 50 minutes at
-78(C, 250ml 0.2M HCl was added and the mixture extracted with diethyl ether. The
organic fractions were washed with brine and dried (MgSO4). Filtration followed by
rotary evaporation yielded a green syrup. Column chromatography (4" x 7", 10% (
20% ether/hexanes yielded a benzaldehyde (a compound of Formula 13), as a white
solid (16.3g, 71%). 1H NMR (600 MHz, CDCl3) (10.47 (s, 1 H), 7.66 (d, J= 7.3 Hz,
2 H), 7.45 (t, J = 7.0 Hz, 1 H), 7.40 (t, J = 7.3 Hz, 2 H), 6.69 (s, 2 H), 3.64 (s, 3 H),
2.55 (s, 6 H), 1.01 (s, 9 H).
Preparation of a Compound of Formula (14) from compounds of Formulas (10) and
(13)
-
To a stirred solution of 3-isopropyl-4-triisopropylsilyoxy-bromobenzene (a
compound of Formula 10) (11.0g, 29.6mmol) in anhydrous THF (125ml) at -78(C
was added solution of butyllithium in hexanes (2.5M, 32.3mmol). This mixture was
transferred via cannula to a stirred suspension of cerous chloride (7.96g, 32.3mmol) in
anhydrous THF (150ml) at -78(C. After stirring 30 min. at -78(C, the benzaldehyde
(a compound of Formula 13) (9.21g, 26.9mmol) in anhydrous THF (25ml) was added
via cannula. After a further 90 min. at -78(C, dilute HCl (170ml, 0.3M) was added
and the mixture extracted with diethyl ether. Combined organic fractions were
washed with a mixture of brine and saturated NaHCO3, then dried (MgSO4) and
filtered through a Celite plug. Removal of solvent by rotary evaporation yielded a
benzyl alcohol (a compound of Formula 14) as a yellow syrup (20.7g) which was used
directly in the next reaction.
Preparation of a Compound of Formula III
-
To a stirred solution of the crude benzyl alcohol (a compound of Formula 14)
(20.6g) and anhydrous methanol (10.9ml, 269mmol) in anhydrous methylene chloride
(400ml) at -45(C (dry ice/acetonitrile slush) was added TFA (10.4ml, 135mmol).
After stirring 50 min. at -45(C, reaction was quenched with 230 ml of a mixture of
brine, saturated NaHCO3 and water (9:9:4). Layers were separated and the aqueous
phase extracted with diethyl ether. Combined organic fractions were dried (MgSO4)
filtered through Celite and solvent removed by rotary evaporation to yield an orange
oil (20.5g). Column chromatography (silica gel, 4" x 7", 0% (15% ether/hexanes)
yielded a methyl ether (a compound of Formula III) as a yellow syrup (14.8g, 77%
yield from the benzaldehyde, a compound of Formula 13). 1H NMR (600 MHz,
CDCl3) (7.70 (d, J= 6.6 Hz, 2 H), 7.43 (t, J= 7.0 Hz, 1 H), 7.39 (t, J= 7.1 Hz, 2 H),
7.04 (s, 1 H), 6.78 (d, J= 8.1 Hz, 1 H), 6.67 (s, 2 H), 6.64 (d, J= 8.4 Hz, 1 H), 5.69
(s, 1 H), 3.62 (s, 3 H), 3.31 (app. s, 4 H), 2.17 (s, 6 H), 1.28 (heptet, J = 7.5 Hz, 3 H),
1.13 (d, J = 7.0 Hz, 3 H), 1.11 (d, J = 3.7 Hz, 3 H), 1.09 (d, J = 7.3 Hz, 18 H), 1.00 (s,
9 H).
Preparation of a Compound of Formula (15)
-
To a stirred solution of the methyl ether (a compound of Formula III) (4.03g,
6.2mmol) in anhydrous THF (60ml) was added triethylamine trihydrofluoride
(2.02ml, 12.4mmol). After 45 min., reaction mixture was partitioned between
saturated NaHCO3 and diethyl ether. The aqueous layer was further extracted with
ether and combined organic fractions were dried (MgSO4), then filtered through a
Celite plug, and solvent was removed by rotary evaporation to yield a yellow syrup
(4.1 g) which crystallized upon further standing. Recrystallization, first from hexanes,
then 1:3 ether:hexanes yielded a phenol (a compound of Formula 15), as a white solid
(2.40g,85%). 1H NMR (300 MHz, CDCl3) (7.15 (s, 1 H), 6.73 (dd, J= 8.1, 1.2 Hz,
1 H), 6.64 (d, J= 8.3 Hz, 1 H), 6.51 (s, 2 H), 5.70 (s, 1 H), 4.68 (br. s, 1 H), 3.32 (app.
s, 4 H), 2.20 (s, 6 H), 1.25 (heptet, J = 7.3 Hz, 3 H), 1.15 (app. t, J = 6.8 Hz, 6 H),
1.08 (d, J= 7.2 Hz, 18 H).
Preparation of Compound (16-1)
-
To a stirred solution of the phenol (a compound of Formula 15) (2.78g,
6.08mmol) in anhydrous DMF (10ml), was added methyl bromoacetate (864µL,
9.13mmol) and potassium carbonate (1.69g, 12.2mmol). Reaction was quenched after
6 hr. by slow addition of dilute HCl (1M, 30ml). The mixture was diluted with 150ml
water and extracted with diethyl ether. Combined organic fractions were washed
twice with brine, then dried (MgSO4) filtered through a Celite plug and solvent
removed by rotary evaporation to yield a methyl ether as a pale yellow oil (3.7g,
100%). 1H NMR (300 MHz, CDCl3) (7.15 (d, J= 0.9 Hz, 1 H), 6.71 (dd, J= 8.3, 1.6
Hz, 1 H), 6.63 (d, J= 8.4 Hz, 1 H), 6.57 (s, 2 H), 5.70 (s, 1 H), 4.63 (s, 2 H), 3.82 (s, 3
H), 3.32 (app. s, 4 H), 2.22 (s, 6 H), 1.26 (heptet, J= 7.3 Hz, 3 H), 1.15 (app. t, J= 6.5
Hz, 6 H), 1.09 (d, J= 7.2 Hz, 18 H).
-
To a stirred, solution of the methyl ether (2.0g, 3.8mmol), allyltimethylsilane
(15ml, 95mmol) in anhydrous methylene chloride (50ml) cooled to -45(C (dry
ice/acetonitrile slush), was added TFA (2.9ml, 38mmol). After stirring 15 min. at -45
(C, reaction was quenched with 50ml of saturated NaHCO3. The aqueous layer was
extracted with diethyl ether and combined organic fractions were dried (MgSO4),
filtered through a Celite plug and solvent removed by rotary evaporation to yield a
cloudy tan syrup (1.92g). Column chromatography (silica, 1.5" x 5", 10%
EtOAc/hexanes yielded the alkene, Compound (16-1) (1.68g, 80%).
Preparation of Compound (18)
-
To a stirred solution of Compound (16-1), (1.68g, 3.12mmol) in anhydrous
THF (40ml), cooled on an ice bath, was added borane (1.0M in THF, 3.12mmol).
Reaction was allowed to warm to room temperature and stirred 4.5 hr. Additional
borane (0.6mmol) was added to drive hydroboration to completion, and reaction was
stirred an additional 3 hr. Hydrogen peroxide (318µl, 3.12mmol) and NaOH (1 M,
0.94mmol) diluted in water (3ml) were then added, and the reaction was stirred 20
min. Saturated NH4Cl was then added, layers separated, and the aqueous phase was
extracted with diethyl ether. Combined organic fractions were washed with brine and
dried (MgSO4) filtered through Celite and solvent removed by rotary evaporation to
yield the crude product (1.81g). Column chromatography (silica, 1" x 5", 20% (35%
EtOAc/hexanes) yielded the alcohol, Compound (18) (836mg, 48%). 1H NMR (600
MHz, CDCl3) (7.00 (s, 1 H), 6.70 (d, J = 8.4 Hz, 1 H), 6.63 (d, J= 8.1 Hz, 1 H), 6.53
(br. s, 2 H), 4.60 (s, 2 H), 4.40 (t, J = 7.7 Hz, 1 H), 3.81 (s, 3 H), 3.66 (heptet, J= 6.4
Hz, 1 H), 3.33 (t, J= 6.8 Hz, 2 H), 2.12 (br. s, 6 H), 2.37-2.32 (m, 1 H), 2.06-1.99 (m,
1 H), 1.65-1.58 (m, 1 H), 1.45-1.38 (m, 1 H), 1.28 (heptet, J= 7.5 Hz, 3 H), 1.15 (d, J
= 6.9 Hz, 3 H), 1.12 (d, J= 7.0 Hz, 3 H), 1.09 (d, J= 7.3 Hz, 18 H).
Preparation of Compound (19)
-
To a stirred suspension of Compound (18), (655mg, 116mmol), potassium
bromide (14mg, 0.12mmol) and TEMPO in methylene chloride (10ml), cooled to 0(C,
was slowly added a mixture of sodium hypochlorite solution (1.87ml, 1.39mmol) and
saturated sodium bicarbonate (2ml). After 40 min., 10ml of 10% HCl containing
potassium iodide (125mg) was added and the aqueous layer extracted with diethyl
ether. Combined organic phases were washed with 10% sodium thiosulfate and 1:1
brine:water, then dried (MgSO4) filtered through Celite and solvent removed by
rotary evaporation to yield the aldehyde, Compound (19), as a yellow oil (615mg,
96%). 1H NMR (600 MHz, CDCl3) (9.71 (s, 1 H), 6.99 (s, 1 H), 6.71 (d, J= 6.2 Hz,
1 H), 6.64 (d, J= 7.1 Hz, 1 H), 6.55 (s, 2 H), 4.60 (s, 2 H), 4.42-4.40 (m, 1 H), 3.82 (s,
3 H), 3.33 (heptet, J= 7.0 Hz, 1 H), 2.64-2.58 (m, 1 H), 2.47-2.41 (m, 1 H), 2.35-2.29
(m, 2 H), 2.13 (br. s, 6 H), 1.28 (heptet, J= 7.4 Hz, 3 H), 1.16 (d, J= 7.0Hz, 3 H),
1.12 (d, J = 7.0 Hz, 3 H), 1.09 (d, J= 7.3 Hz, 18 H).
Preparation of Compound (21)
-
To a stirred suspension of commercially available 8-bromooctanoic acid,
Compound (20), (1.27g, 5.69mmol) and HBTU (2.16g, 5.69mmol) in anhydrous
methylene chloride (20ml) was added N-methylbutylamine (673µl, 5.69mmol) and
triethylamine (1590µl, 11.4mmol). After 4 hr., brine was added and the aqueous
phase extracted with diethyl ether. The combined organic fractions were washed with
1M HCl (3 x 20ml), saturated NaHCO3 (2 x 20ml) and brine, then dried (MgSO4),
filtered through Celite and solvent removed by rotary evaporation. Column
chromatography (silica gel, 1.5" x 5", 20% (30% EtOAc/Hexanes) yielded the
bromooctamide, Compound (21), as a colorless oil (1.219g, 76%). 1H NMR (600
MHz, CDCl3) (3.40 (t, J= 7.0 Hz, 2 H), 3.36 (t, J= 7.3 Hz, 1 H), 3.25 (t, J= 7.3 Hz,
1 H), 2.97 (s, 1.5 H), 2.91 (s, 1.5 H), 2.29 (app. q, J = 7.4 Hz, 2 H), 1.86 (quintet, J=
7.1 Hz, 2 H), 1.66-1.62 (m, 2 H), 1.54 (quintet, J= 7.4 Hz, 1 H), 1.49 (quintet, J =
7.5 Hz, 1 H), 1.46-1.42 (m, 2 H), 1.36-1.28 (m, 6 H), 0.96 (t, J= 7.3 Hz, 1.5 H), 0.92
(t,J = 7.3Hz, 1.5 H).
Preparation of Compound (22)
-
To undiluted Compound (21) (117mg , 0.400mmol) was added
triphenylphosphine (105mg, 0.400mmol). The mixture was stirred at 125(C for 22hr.,
then cooled and dissolved in anhydrous THF (4ml). To this solution was added
potassium tert-butoxide (45mg, 0.400mmol) and then aldehyde 19 (200mg,
0.36mmol), dissolved in anhydrous THF. Reaction was heated to reflux for 7hr., then
quenched with saturated NH4Cl and the aqueous phase extracted with diethyl ether.
Combined organic fractions were washed with brine, dried (MgSO4) filtered through a
Celite pad and solvent removed by rotary evaporation. This procedure was repeated
once and the crude products chromotographed twice under different conditions (silica
gel, 1" x 6", 10% (35% EtOAc/Hexanes) (silica gel, 0.5" x 7". 2.5% (20%
EtOAc/Methylene chloride) to yield the alkene, Compound (22), as a colorless oil
(79mg, 13%).
Preparation of Compound (23)
-
To a stirred solution of Compound (22) (29mg, 0.039mmol) in absolute
ethanol was added palladium on carbon (spatula tip). The flask was purged with
hydrogen gas (balloon) and stirred 24hr. under balloon pressure. Filtration through
Celite and rotary evaporation yielded the ester, Compound (23) (25.5mg, 87%). 1H
NMR (600 MHz, CDCl3) (6.98 (s, 1 H), 6.70 (d, J= 8.4 Hz, 1 H), 6.62 (d, J= 8.4 Hz,
1 H), 6.53 (br. s, 2 H), 4.60 (s, 2 H), 4.37-4.34 (m, 1 H), 3.81 (s, 3 H), 3.37-3.30 (m, 2
H), 3.25 (t, J = 7.5 Hz, 1 H), 2.96 (s, 1.5 H), 2.90 (s, 1.5 H), 2.30-2.26 (m, 2 H),
2.25-2.21 (m, 1 H), 2.04 (br. s, 6 H), 1.95-1.89 (m, 1 H), 1.64-1.60 (m, 2 H),
1.57-1.52 (m, 1 H), 1.51-1.46 (m, 1 H), 1.35-1.24 (m, 19 H), 1.15 (d, J= 7.0 Hz, 3 H),
1.12 (d, J= 7.0 Hz, 3 H), 1.09 (d, J= 7.3 Hz, 18 H), 0.96-0.91 (m, 3H).
Preparation of Compound (24)
-
To a stirred solution of Compound (23) (25.5mg, 0.034mmol) in anhydrous
THF (500µl) was added triethylamine trihydrofluoride (552µl, 3.4mmol). After 18hr.,
reaction was quenched with potassium carbonate (700mg, 5.1mmol) and water (3ml),
then extracted with ethyl acetate. Combined organic fractions were dried (MgSO4),
filtered through Celite and solvent removed by rotary evaporation. Column
chromatography (silica gel, 0.5" x 6". 10% EtOAc/Hexanes (20% EtOAc/1%
MeOH/Hexanes) yielded the phenol, Compound (24), (12mg, 59%) and unreacted
starting material (9.5mg).
Preparation of Compound (I-1)
-
To a stirred solution of Compound (24) (12mg, 0.020mmol) in methanol (1ml)
was added lithium hydroxide monohydrate (2mg, 0.05mmol) and water (2µl,
0.11mmol). After 16hr., saturated NH4Cl and two drops of 1M HCl were added and
the mixture was extracted with ethyl acetate. The combined organic fractions were
dried (MgSO4) filtered through Celite and solvent removed by rotary evaporation to
yield the oxyacetic acid, Compound (I-5) (6mg, 52%). 1H NMR (600 MHz, CDCl3) (
7.00 (s, 1 H), 6.75 (d, J= 7.3 Hz, 1 H), 6.61 (d, J= 8.1 Hz, 1 H), 6.56 (br. s, 2 H),
4.62 (s, 2 H), 4.38-4.35 (m, 1 H), 3.37 (t, J= 7.5 Hz, 1 H), 3.26 (t, J= 7.5 Hz, 1 H),
3.16 (heptet, J= 7.1 Hz, 1 H), 2.98 (s, 1.5 H), 2.93 (s, 1.5 H), 2.36-2.22 (m, 2 H),
2.21-2.15 (m, 1 H), 2.10 (br. s, 6 H), 2.05-1.96 (m, 1 H), 1.63-1.57 (m, 2 H),
1.56-1.52 (m, 1 H), 1.52-1.47 (m, 1 H), 1.37-1.29(m, 4 H), 1.28-1.23 (m, 4 H), 1.21
(app. d, J= 7.0 Hz, 6 H), 1.18 (app. d, J= 7.0 Hz, 4 H), 1.15-1.11 (m, 4 H), 0.95 (t, J
= 7.5 Hz, 1.5 H), 0.92 (t, J= 7.3 Hz, 1.5 H).
-
HRMS exact mass calcd for C36H55NO5: 581.4080, found: 581.4082.
EXAMPLE 12
Preparation of Compound (I-2), a Compound of Formula I
Preparation of Compound (25)
-
To a stirred solution of Compound (22) (50mg, 0.067mmol, from Example 11)
in anhydrous THF (1ml) was added triethylamine trihydrofluoride (1.09ml,
6.67mmol). After 13hr., reaction was quenched with potassium carbonate (1.28g) and
water (5ml) and extracted with chloroform. Combined organic fractions were dried
(MgSO4) filtered through Celite, and solvent was removed by rotary evaporation.
Column chromatography (silica, 0.5" x 7", 10% EtOAc/CH2Cl2 (15% EtOAc/3%
MeOH/CH2Cl2) yielded the phenol, Compound (25) (33mg, 83%). 1H NMR (600
MHz, CDCl3) (6.99 (s, 1H), 6.74 (d, J= 7.3 Hz, 1 H), 6.67 (d, J= 8.1 Hz, 1 H), 6.52
(s, 2 H), 6.05 (br. s, 1 H), 5.40-5.35 (m, 2 H), 4.59 (s, 2 H), 4.36 (t, J= 7.3 Hz, 1 H),
3.81 (s, 3 H), 3.37 (t, J= 7.5 Hz, 1 H), 3.26-3.19 (m, 2 H), 2.96 (s, 1.5 H), 2.92 (s, 1.5
H), 2.40-2.32 (m, 1 H), 2.28 (app. q, J= 8.1 Hz, 2 H), 2.15 (br. s, 6 H), 2.13-2.09 (m,
1 H), 1.92-1.87 (m, 1 H), 1.86-1.81 (m, 2 H), 1.66-1.47 (m, 3 H), 1.35-1.27 (m, 4 H),
1.26-1.22 (m, 2 H), 1.21 (d, J= 7.0 Hz, 3 H), 1.18 (d, J= 6.6 Hz, 3 H), 1.16-1.14 (m,
1 H), 1.09 (app. d, J = 7.3 Hz, 3 H), 0.95 (t, J= 7.5 Hz, 1.5 H), 0.92 (t, J = 7.3 Hz, 1.5
H).
Preparation of Compound (I-2)
-
To a stirred solution of Compound (25) (10mg, 0.017mmol) in methanol was
added lithium hydroxide monohydrate (3mg, 0.07mmol) and water (1.7µl,
0.093mmol). After 26hr., solvent was evaporated and the residue resuspended in
saturated NH4Cl and a drop of 1M HCl., which was extracted with ethyl acetate.
Combined organic fractions were dried (MgSO4), filtered through Celite, and solvent
was removed by rotary evaporation to yield the oxyacetic acid, Compound (I-2) (7mg,
71%). 1H NMR (600 MHz, CDCl3) (7.01 (s, 1 H), 6.74 (d, J= 7.7 Hz, 1 H), 6.62 (d,
J= 8.4 Hz, 1 H), 6.56 (s, 2 H), 5.46-5.41 (m, 1 H), 5.36-5.32 (m, 1 H), 4.63 (s, 2 H),
4.3 8-4.3 6 (m, 1 H), 3 .38 (t, J = 7.5 Hz, 1 H), 3.26 (t, J = 7.3 Hz, 1 H), 3.17 (heptet, J
= 6.6 Hz, 1 H), 2.99 (s, 1.5 H), 2.94 (s, 1.5 H), 2.38-2.30 (m, 2 H), 2.10 (br. s, 6 H),
2.06-1.95 (m, 2 H), 1.94-1.86 (m, 1 H), 1.75-1.68 (m, 2 H), 1.53-1.47 (m, 1 H),
1.36-1.28 (m, 3 H), 1.26 (app. s, 1 H), 1.21-1.18 (m, 9 H), 1.16-1.11 (m, 3 H),
0.96-0.91 (m, 3 H).
-
HRMS exact mass calculated for C36H53NO5: 579.3924, found: 579.3924.
EXAMPLE 13
Preparation of Compound (I-3), a Compound of Formula I
Preparation of Compound (26)
-
To a stirred solution of the phenol (compound of Formula 15 from Example
11) (2.32g, 4.93mmol) in anhydrous DMF (100ml) was added cesium carbonate
(3.21g, 9.86mmol) and ethyl bromoacetate (819µl, 7.39mmol). Reaction was stirred 3
hr., then quenched with 150ml saturated NH4Cl, then diluted with water (300ml).
Extraction with diethyl ether, followed by drying (MgSO4) filtration and rotary
evaporation yielded an oil (3.0g). This contained some residual DMF, which was
removed by partitioning the residue between ether and water to yield 2.82g of a
methyl ether. 1H NMR (600 MHz, CDCl3) (7.14 (s, 1 H), 6.71 (d, J= 8.0 Hz, 1 H),
6.63 (d, J= 8.4 Hz, 1 H), 6.58 (s, 2 H), 5.70 (s, 1 H), 4.61 (s, 2 H), 4.29 (q, J= 7.1
Hz, 2 H), 3.32 (app. s, 4 H), 2.22 (s, 6 H), 1.32-1.25 (m, 6 H), 1.16 (d, J= 7.0 Hz, 3
H), 1.15 (d, J= 6.6 Hz, 3 H), 1.09 (d, J= 7.3 Hz, 18 H).
-
To a stirred solution of the methyl ether, (1.05g, 1.93mmol) and
allyltrimethylsilane (13.2ml, 83.3mmol) in anhydrous methylene chloride (50ml),
chilled to -45(C (dry ice/acetonitrile slush), was added TFA (2.1ml, 28mmol). After
20 min. at -45(C, cold bath was removed, and the reaction was quenched with 50ml
saturated NaHCO3. The aqueous phase was extracted with diethyl ether, and the
combined organic fractions were dried (MgSO4), filtered, and solvent was removed by
rotary evaporation. Column chromatography (silica, 1.5" x 7", 7% (12%
EtOAc/Hexanes) yielded the alkene, Compound (26) as a colorless oil (795mg, 75%).
1H NMR (600 MHz, CDCl3) (7.01 (s, 1 H), 6.70 (d, J = 8.1 Hz, 1 H), 6.63 (d, J= 8.4
Hz, 1 H), 6.53 (s, 2 H), 5.72-5.68 (m, 1 H), 5.07 (d, J= 16.9 Hz, 1 H), 4.92 (d, J=
10.3 Hz, 1 H), 4.5 8 (s, 2 H), 4.49 (t, J= 7.9 Hz, 1 H), 4.28 (q, J= 7.1 Hz, 2 H), 3.33
(heptet, J= 7.0 Hz, 1 H), 3 .04-2.99 (m, 1 H), 2.74-2.69 (m, 1 H), 2.12 (br. s, 6 H),
1.30-1.25 (m, 6 H), 1.15 (d, J= 7.0 Hz, 3 H), 1.12 (d, J= 7.0 Hz, 3 H), 1.09 (d, J=
7.3 Hz, 18 H).
Preparation of Compound (I-3)
-
To a stirred solution of Compound (26) (100mg, 0.181mmol) in anhydrous
THF (5ml) was added triethylamine trihydrofluoride (2ml, 12mmol). After 39hr.,
reaction was quenched with 20ml saturated NaHCO3 and extracted with diethyl ether.
Combined organic fractions were dried (MgSO4), filtered, and solvent was removed
by rotary evaporation. Column chromatography (silica, 0.5" x 5 ", 10% (14%
EtOAc/Hexanes) yielded a phenol (57mg, 79%).
-
To a stirred solution of the phenol (57mg, 0.14mmol) in methanol (3ml) was
added lithium hydroxide monohydrate (13mg, 0.32mmol) and water (14µl,
0.79mmol). After 13hr., solvent was removed by rotary evaporation, and the residue
was resuspended in saturated NH4Cl, plus two drops of 1M HCl., and extracted with
chloroform. Combined organic fractions were dried (MgSO4), filtered, and solvent
was removed by rotary evaporation to yield the oxyacetic acid, Compound (I-3), as a
colorless oil (42mg, 79%). 1H NMR (300 MHz, CDCl3) (7.02 (s, 1 H), 6.75 (dd, J=
8.1, 1.2 Hz, 1 H), 6.61 (d, J = 8.3 Hz, 1 H), 6.55 (s, 2 H), 5.77-5.65 (m, 1 H), 5.08 (d,
J= 16.9 Hz, 1 H), 4.93 (d, J= 10.2 Hz, 1 H), 4.64 (s, 2 H), 4.49 (t, J= 7.9 Hz, 1 H),
3.16 (heptet, J= 6.9 Hz, 1 H), 3.06-2.98 (m, 1 H), 2.78-2.68 (m, 1 H), 2.14 (br. s, 6
H), 1.19 (app. t, J = 7.7 Hz, 6 H).
-
HRMS exact mass calcd for C23H28O4: 368.1988, found: 368.1994.
EXAMPLE 14
Preparation of Compound (1-4), a Compound of Formula I
Preparation of Compound (27)
-
To a stirred solution of Compound (26) (320mg, 0.579mmol, from Example
13) in anhydrous THF (5ml) was added borane (1M in THF, 790mmol), after 20hr., a
mixture of sodium hydroxide (0.29mmol) and hydrogen peroxide (30%, 60µl,
0.579mmol) was added. After 1hr., reaction was quenched with saturated NH4Cl and
the aqueous phase extracted with diethyl ether. Combined organic fractions were dried
(MgSO4), filtered, and solvent was removed by rotary evaporation. Column
chromatography (silica, 1" x 7 ", 15% (25% EtOAc/Hexanes) yielded the alcohol,
Compound (27), as a colorless oil (170mg, 51%).
Preparation of Compound (28)
-
To a stirred solution of Compound (27) (67mg, 0.12mmol) in anhydrous THF
(4ml) was added triethylamine trihydrofluoride (1.9ml, 12mmol). Reaction was
stirred 15hr. then quenched with 20ml saturated NaHCO3 and extracted with diethyl
ether.
Combined organic fractions were dried (MgSO4), filtered through Celite, and solvent
was removed by rotary evaporation. Column chromatography (silica, 0.5" x 5 ", 30%
EtOAc (40% EtOAc/2% AcOH/Hexanes) yielded the phenol, Compound (28) (33mg,
66%). 1H NMR (300MHz, CDCl3) (6.98 (s, 1 H), 6.74 (dd, J = 8.2, 1.3 Hz, 1 H),
6.60 (d, J= 8.2 Hz, 1 H), 6.54 (s, 2 H), 4.58 (s, 2 H), 4.41-4.36 (m, 1 H), 4.28 (q, J=
7.1 Hz, 2 H), 3.66 (t, J = 6.4 Hz, 2 H), 3.17 (heptet, J= 6.9 Hz, 1 H), 2.39-2.27 (m, 1
H), 2.13-1.97 (br. m, 7 H), 1.68-1.56 (m, 1 H), 1.45-1.35 (m, 1 H), 1.29 (t, J= 7.2 Hz,
3 H), 1.19 (d, J= 7.1 Hz, 3 H), 1.17 (d, J= 7.0 Hz, 3 H).
Preparation of Compound (I-4)
-
To a stirred solution of Compound (28) (33mg, 0.080mmol) in methanol (3ml)
was added lithium hydroxide monohydrate (7.4mg, 0.18mmol) and water (8µl,
0.4mmol). Reaction was stirred 13hr., then solvent was removed by rotary
evaporation. The residue was resuspended in 1/2-saturated NH4Cl and extracted with
chloroform. Combined organic fractions were dried (MgSO4), filtered through Celite,
and solvent was removed by rotary evaporation to yield the oxyacetic acid, Compound
(I-4), as a white solid (22mg, 71%). 1H NMR (300MHz, CDCl3) (6.97 (s, 1 H), 6.73
(d, J= 7.6 Hz, 1 H), 6.64 (d, J = 8.2 Hz, 1 H), 6.57 (s, 2 H), 4.58 (s, 2 H), 4.43-4.38
(m, 1 H), 3.59 (t, J= 6.4 Hz, 2 H), 3.23 (heptet, J= 6.9 Hz, 1 H), 2.39-2.27 (m, 1 H),
2.25-1.96 (br. m, 7 H), 1.63-1.54 (m, 1 H), 1.42-1.32 (m, 1 H), 1.17 (d, J= 7.2 Hz, 3
H), 1.13 (d, J= 7.3 Hz, 3 H).
-
HRMS exact mass calcd for C23H30O5: 386.2093, found: 386.2096.
EXAMPLE 15
Preparation of Compound (I-5), a Compound of Formula I
Preparation of Compound (29)
-
To a stirred solution of the phenol (compound of Formula 15 from Example
11) (2.78g, 6.08mmol) in anhydrous DMF (10ml), was added methyl bromoacetate
(864/µL, 9.13mmol) and potassium carbonate (1.69g, 12.2mmol). Reaction was
quenched after 6 hr. by slow addition of dilute HCl (1M, 30ml). Mixture was diluted
with 150ml water and extracted with diethyl ether. Combined organic fractions were
washed twice with brine, then dried (MgSO4), filtered through a Celite plug and
solvent removed by rotary evaporation to yield the methyl ether, Compound (29) as a
pale yellow oil (3.7g, 100%). 1H NMR (300 MHz, CDCl3) (7.15 (d, J= 0.9 Hz, 1
H), 6.71 (dd, J= 8.3, 1.6 Hz, 1 H), 6.63 (d, J= 8.4 Hz, 1 H), 6.57 (s, 2 H), 5.70 (s, 1
H), 4.63 (s, 2 H), 3.82 (s, 3 H), 3.32 (app. s, 4 H), 2.22 (s, 6 H), 1.26 (heptet, J= 7.3
Hz, 3 H), 1.15 (app. t, J= 6.5 Hz, 6 H), 1.09 (d, J= 7.2 Hz, 18 H).
Preparation of Compound (30)
-
To a stirred solution of Compound (29) (150mg, 0.28mmol) and
1,3-dimethyoxybenzene (470mg, 3.4mmol) in anhydrous methylene chloride (7ml),
chilled to -45(C, was added TFA (220/µl, 2.8mmol). After stirring 90min, while
allowing gradual warming, reaction was quenched with 13ml saturated NaHCO3 and
the aqueous phase was extracted with diethyl ether. Combined organic fractions were
dried (MgSO4), filtered through Celite, and solvent was removed by rotary
evaporation to yield Compound (30) as a colorless oil (586mg), which was used
without further purification.
Preparation of Compound (31)
-
To a stirred solution of Compound (30) (crude, approx. 0.26mmol) in
anhydrous THF (5ml) was added triethylamine trihydrofluoride (4.6ml, 28mmol).
After 10hr., reaction was quenched with 15ml 4M NaOH. The aqueous phase was
extracted with diethyl ether and the combined organic fractions were dried (MgSO4),
filtered through Celite, and solvent was removed by rotary evaporation. Column
chromatography (silica, 1" x 6", 15% (35% EtOAc/Hexanes) yielded Compound (31)
(74mg, 55% yield from Compound (29)). 1H NMR (300MHz, CDCl3) (6.81 (s, 1 H),
6.74 (d, J= 8.4 Hz, 1 H), 6.58 (s, 2 H), 6.54 (s, 2 H), 6.47 (d, J= 2.1 Hz, 1 H), 6.36
(dd, J= 8.5, 2.1 Hz, 1 H), 5.90 (s, 1 H), 4.61 (s, 2 H), 3.81 (s, 3 H), 3.79 (s, 3 H), 3.66
(s, 3 H), 3.13 (heptet, J = 6.9 Hz, 1 H), 1.97 (s, 6 H), 1.13 (d, J = 6.9 Hz, 6 H).
Preparation of Compound (I-5)
-
To a stirred solution of Compound (31) (74mg, 0.15mmol) in methanol was
added lithium hydroxide monohydrate (14mg, 0.34mmol) and water (15µl,
0.85mmol). After 7hr., solvent was removed by rotary evaporation. The residue was
resuspended in 4ml saturated NH4Cl + 3ml water + 1ml 1M HCl and extracted with
chloroform. Combined organic fractions were dried (MgSO4) filtered through Celite,
and solvent was removed by rotary evaporation to yield an oxyacetic acid, Compound
(I-5), as a white semi-solid (77mg, 100%). 1H NMR (300MHz, CDCl3) (6.81 (s, 1
H), 6.74 (d, J = 8.5 Hz, 1 H), 6.58 (s, 2 H), 6.56 (s, 2 H), 6.47 (d, J = 2.0 Hz, 1 H),
6.36 (dd, J= 8.4, 2.0 Hz, 1 H), 5.90 (s, 1 H), 4.65 (s, 2 H), 3.79 (s, 3 H), 3.66 (s, 3 H),
3.13 (heptet, J= 6.8 Hz, 1 H), 1.98 (s, 6 H), 1.13 (d, J= 6.8 Hz, 6 H).
-
HRMS exact mass calcd for C28H32O6: 464.2199, found: 464.2192.
EXAMPLE 16
-
This example illustrates the preparation of a representative pharmaceutical
formulation for oral administration containing an active compound of Formula_I, e.g.,
ethoxy-2,6-dimethyl-4'-hydroxy -3'-(1-methylethyl)diphenylmethane-4-oxyacetic acid.
Ingredients | Quantity per Tablet, mgs. |
Active Compound | 200 |
Lactose, spraydried | 148 |
Magnesium stearate | 2 |
The above ingredients are mixed and introduced into a hard-shell gelatin capsule.
-
Other compounds of Formula_I can be used as the active compound in the
preparation of the orally administrable formulations of this example.
EXAMPLE 17
-
This example illustrates the preparation of another representative
pharmaceutical formulation for oral administration containing a compound of
Formula_I, e.g., ethoxy-2,6-dimethyl-4'-hydroxy
-3'-(1-methylethyl)diphenylmethane-4-oxyacetic acid.
Ingredients | Quantity per Tablet, mgs. |
Active Compound | 400 |
Cornstarch | 50 |
Lactose | 145 |
Magnesium stearate | 5 |
The above ingredients are mixed intimately and pressed into single scored tablets.
-
Other compounds of Formula_I can be used as the active compound in the
preparation of the orally administrable formulations of this example.
EXAMPLE 18
-
This example illustrates the preparation of a representative pharmaceutical
formulation containing a compound of Formula_I, e.g.,
ethoxy-2,6-dimethyl-4'-hydroxy-3'-(1-methylethyl)diphenylmethane-4-oxyacetic acid.
-
An oral suspension is prepared having the following composition.
Ingredients | Quantity |
Active Compound | 1.0 g |
Fumaric acid | 0.5 g |
Sodium chloride | 2.0 g |
Methyl paraben | 0.1 g |
Granulated sugar | 25.5 g |
Sorbitol (70% solution) | 12.85 g |
Veegum K (Vanderbilt Co.) | 1.0 g |
Flavoring | 0.035 ml |
Colorings | 0.5 mg |
Distilled water | q.s. to 100 ml |
-
Other compounds of Formula_I can be used as the active compound in the
preparation of the orally administrable formulations of this example.
EXAMPLE 19
-
This example illustrates the preparation of a representative pharmaceutical
formulation for oral administration containing an active compound of Formula_I, e.g.,
ethoxy-2,6-dimethyl-4'-hydroxy-3'-(1-methylethyl)diphenylmethane-4-oxyacetic acid.
-
An injectable preparation buffered to a pH of 4 is prepared having the
following composition:
Ingredients | Quantity |
Active Compound | 0.2 g |
Sodium Acetate Buffer Solution (0.4 M) | 2.0 ml |
HCl (IN) | q.s. to pH 4 |
Water (distilled, sterile) | q.s. to 20 ml |
-
Other compounds of Formula_I can be used as the active compound in the
preparation of the injectable formulations of this example.
EXAMPLE 20
-
This example illustrates the preparation of a representative pharmaceutical
formulation for topical application containing a compound of Formula_I, e.g.,
ethoxy-2,6-dimethyl-4'-hydroxy-3'-(1-methylethyl)diphenylmethane-4-oxyacetic acid.
Ingredients | grams |
Active compound | 0.2-10 |
Span 60 | 2 |
Tween 60 | 2 |
Mineral oil | 5 |
Petrolatum | 10 |
Methyl paraben | 0.15 |
Propyl paraben | 0.05 |
BHA (butylated hydroxy anisole) | 0.01 |
Water | q.s. to 100 |
-
All of the above ingredients, except water, are combined and heated to 60°C
with stirring. A sufficient quantity of water at 60°C is then added with vigorous
stirring to emulsify the ingredients, and water then added q.s.100_g.
-
Other compounds of Formula_I can be used as the active compound in the
preparation of the topical formulations of this example.
EXAMPLE 21
-
This example illustrates the preparation of a representative pharmaceutical
formulation containing a compound of Formula_I, e.g.,
ethoxy-2,6-dimethyl-4'-hydroxy -3'-(1-methylethyl)diphenylmethane-4-oxyacetic acid.
-
A suppository totaling 2.5_grams is prepared having the following
composition:
Ingredients | Quantity |
Active Compound | 500 mg |
Witepsol H-15* | balance |
(*triglycerides of saturated vegetable fatty acid; a product of Riches-Nelson, Inc., New York, NY) |
-
Other compounds of Formula_I can be used as the active compound in the
preparation of the suppository formulations of this example.
EXAMPLE 22
Receptor Binding Assays of TR Ligands
-
To test the ability of synthesized human thyroid receptor (hTR) ligands to bind
to two subtypes of hTR, hTRα and hTRβ, the binding affinity of a TR ligand for a TR
can be assayed using TRs expressed in E. coli and [125I] T3 (radiolabeled
3,5,3'-triiodo-L-thyronine) using the method described by Apriletti et al., Protein
Expression and Purification, 6:363-370 (1995), and by Apriletti et al., J. Biol. Chem.
(1988) which are incorporated herein by reference. The TR binding experiment is
conducted using the recombinant TRs in the presence of the sample to be assayed, 1
nM [125I]T3, and 50(g/ml core histones, in buffer E (400 mM KCI, 200 mM potassium
phosphate, pH 8.0, 0.5 mM EDTA, 1 mM MgCl2, 10% glycerol, 1 mM DTT) in a
volume of 0.21 ml. After incubation overnight at 4°C, 0.2 ml of the incubation
mixture is loaded onto a Quick-Sep Sephadex G-25 column (2.7 x 0.9 cm, 1.7 ml bed
volume) equilibrated with buffer E. The excluded peak of protein-bound [125I]T3 is
eluted with 1 ml of buffer E, collected in a test tube, and counted. Specific T3 binding
is calculated by subtracting nonspecific binding from total binding. The binding
affinity of a ligand for its receptor is defined by a constant termed Kd and can be
calculated using a curve fitting program.
Competition by analogues Compounds (I-1), (I-2) and (I-3)
for [
125
I] T
3
binding to hTRα and β
-
The ability of [125I] T3 and each of Compounds (I-1), (I-2) and (I-3) to compete
for binding to human recombinant TRα and separately human recombinant TRβ was
measured through competition assays. In control experiments, either purified
recombinant hTRα or hTRβ was incubated with [125I] T3 and increasing
concentrations (10-10 M to 10-7M) of unlabelled T3, and the ability of [25I] T3 to
compete with unlabelled T3 for binding to each of the two TR subtypes was measured.
As expected, the unlabelled T3 was able to compete out [125I] T3 in binding to both
hTRα or hTRβ, with Kd values of about 0.069 nM and 0.040 nM respectively. To
test the analogues, either purified recombinant hTRα or hTRβ was incubated with
[125I] T3 and increasing concentrations of unlabelled Compound (I-3) (10-7 M to 10-4
M) or Compound (I1) (10-8 M to 10-5 M) or Compound (I-2) (10-8 M to 10-5 M). The
ability of each analogue to compete with T3 for binding to each of the two TR
subtypes was measured. Unlabelled Compound (I-3) was able to compete with [125I]
T3 for binding to either hTRα or hTRβ, with Kd values of about 138nM and 36 nM
respectively. Unlabelled Compound (I-1) was able to compete with [125I] T3 for
binding to either hTRα or hTRβ, with Kd values of about 77nM and 180nM
respectively. Unlabelled Compound (I-2) was able to compete with [125I] T3 for
binding to either hTRα or hTRβ, with Kd values of about 237nM and 721nM
respectively.
EXAMPLE 23
Cellular Transcription Assay of TR Ligands
Cell Culture, Transfections and Luciferase Assay
-
Cellular transactivation assays can be performed according to the procedure in
Ribeiro R.C. et al. (1996) J. Biol. Chem. 271, 17147-17151. Briefly, HeLa cells are
grown in 15 cm dishes in DME H-21, 4.5 g/L glucose with 10% newborn bovine
serum, 2mM glutamine, 50 units/ml penicillin, and 50 (g/ml streptomycin.
-
For transfections, cells are trypsinized, resuspended in buffer (PBS, 0.1%
glucose), and mixed with a reporter gene construct and with or without the appropriate
thyroid receptor (TR) expression vectors (CMV TR (1, CMV TR (1). One such
reporter gene construct consists of a synthetic TR response element (DR-4) containing
two copies of a direct repeat spaced by four nucleotides (AGGTCA-caggAGGTCA)
cloned into the HindIII site of the pUC 19 polylinker immediately upstream of a
minimal (-32/+45) thymidine kinase promoter linked to luciferase coding sequences.
Another reporter gene construct that can be used consists of the β-galactosidase
coding sequence fused downstream of an actin promoter.
-
Cells in 0.5 ml of buffer (8+/- 2 million cells) are electroporated using a
Bio-Rad gene pulser at 0.35 kvolts and 960 microfarads. After electroporation, cells
are pooled in growth medium (DME H-21 with 10% charcoal-treated, hormone
stripped, newborn bovine serum), plated in 6-well dishes, and are treated with either
vehicle (ethanol), hormone (T3), or analogue (the test ligand). T3 and the test ligand
are used at a range of selected concentrations. After incubation at 37°C for 24 hours,
incubation media is discarded and the cells are detached with 1 ml of
calcium/magnesium-free PBS, 1mM EDTA, prewarmed at 37°C, and transferred to
1.5 ml Eppendorf tubes. Cells are pelleted by centrifugation in a microfuge for 15
seconds at room temperature. The supernatants are aspirated and the pellets are lysed
by addition of 120 (1 of Tris-Cl 0.25 M pH 7.6, 0.1 % Triton. After resuspension by
vortexing for 5-10 sec, the lysates are pelleted by centrifugation in a microfuge for 5
min at room temperature. One hundred (1 of each Eppendorf tube lysate is added to
300 (1 of 25 mM glycylglycine pH 7.8,15 mM MgSO4,4 mM EGTA, 15 mM
potassium phosphate pH 7.8, 1 mM DTT, 2 mM ATP, and 0.2 mM Luciferine. The
light output is measured for 10 sec at room temperature with a luminometer
(Analytical Luminescence Laboratory, MONOLIGHTR 1500).
Transcription Activation of TR by Compounds (I-1) and (I-3)
-
The ability of Compound (I-1) and Compound (I-3) to activate transcription
via each of the two subtypes of hTR was measured by Luciferase assay. In control
experiments, HeLa cells overexpressing either hTRα or hTRβ and containing the
luciferase reporter gene construct were incubated with increasing concentrations (10-12
to 10-7 M) of T3. The ability of T3 to bind to each of the two hTR subtypes, interact
with the TR response element on the reporter gene construct and allow the
downstream promoter to drive the expression of the luciferase protein was measured.
Luciferase elicits a light output than allows for detection of the expressed protein. As
expected, T3 was able to activate transcription and translation of the luciferase gene.
To test analogue Compounds (I-3) and (I-1), HeLa cells overexpressing either hTRα
or hTRβ and containing the luciferase reporter gene construct were incubated with
increasing concentrations of Compound (I-3) (10-10 M to 3x10-5 M) or Compound (I-1)
(10-9 M to 3x10-5 M). Compound (I-3) was able to stimulate transcription and
translation of the luciferase gene through both subtypes of hTR, though to a lesser
degree than T3, indicating that Compound (I-3) may serve as a weak agonist of both
hTRα and hTRβ. Compound (I-1) did not appear to activate hTR-mediated
transcription though was still able to compete with T3, indicating that Compound (I-1)
may serve as an antagonist of both hTRα and hTRβ.
-
To further test analogue Compound (I-1)'s role, HeLa cells containing the
luciferase reporter gene construct and the β-galactosidase reporter gene construct, but
no overexpressed hTRs were incubated with increasing concentrations of Compound
(I-1) (10-6 M to 3x10-5M) and/or 1nM T3. This experiment showed that the luciferase
activity seen at the highest concentration [10-5] of Compound (I-1) occurs even when a
TR-expression construct is not cotransfected into HeLa cells, and subsequently
correlates with toxicity as defined by, for example, a change in cell morphology, or
lifting off of cells from a growth surface, observed in the micromolar range (10µM).
These results indicate that the observed luciferase activity is not due to the
overexpression of thyroid hormone receptors in HeLa cells. In addition, expression of
β-galactosidase was not affected by the presence of 1 nM T3 or increasing
concentrations of Compound (I-1) (10-6 M to 10-5 M), indicating that the decrease in
reporter gene activity observed in the competition experiment between T3 and
Compound (I-1) was not likely to be due to a decrease in cell density.
-
While the present invention has been described with reference to the specific
embodiments thereof, it should be understood by those skilled in the art that various
changes may be made and equivalents may be substituted without departing from the
true spirit and scope of the invention. In addition, many modifications may be made
to adapt a particular situation, material, composition of matter, process, process step
or steps, to the objective, spirit and scope of the present invention. All such
modifications are intended to be within the scope of the claims appended hereto.